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  • 96
    Thermo Fisher cd34
    Significantly decreased colony-forming efficiency of Smad4 mutant keratinocytes. (A and B) Flow cytometry of α6-integrin and <t>CD34</t> double-labeled keratinocytes from dorsal epidermis of wild-type and Smad4 mutant mice at P72. (C–J) Clonal
    Cd34, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 96/100, based on 3850 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore cd34
    Effect of variation in culture methods on phenotype. (A) Effect of FBS content the proportion of the human CDCs expressing markers of CPC (c-Kit), endothelial (CD31, <t>CD34)</t> and mesenchymal (CD90) origin. (B) Flow cytometry dot plot demonstrating the influence of endothelial media and standard cardiac explant media on the proportion of WK rat outgrowth cells expressing the CPC marker c-Kit.
    Cd34, supplied by Millipore, used in various techniques. Bioz Stars score: 95/100, based on 515 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Becton Dickinson cd34
    FACS isolation of <t>CD34</t> + cells infected with lentiviral vectors. Purified CD34 + cells (3 × 10 6 ) were infected with lentiviral vectors (MOI = 3) by centrifugation (4 h at 2,500 rpm) in a Beckman GPR centrifuge. Cells were incubated for 72 h in IMDM supplemented with 10% FBS and 30 μl of cytokine cocktail StemSpan CC100. Cells were incubated with a PE-conjugated human-specific MAb against the CD34 marker, and cell sorting was performed with a FACSVantage flow cytometer. An additional cell sample was incubated with a mouse immunoglobulin G γ isotype control antibody to set compensation and gates for FACS. The R2 gate in each panel represents the gate set to isolate CD34 + GFP + ). The percentage of CD34 + GFP + cells is displayed in the upper right panel. Mock-transduced CD34 + cells were isolated only on the basis of CD34 + expression and are represented by the R3 gate. LVs used in each transduction are as follows: GFP, HR′CMV-GFP; Tax1, HR′CMV-Tax1/GFP; Tax1(−), HR′CMV-Tax1(−)/GFP; Tax2, HR′CMV-Tax2/GFP; and Vpr, HR′CMV-Vpr/GFP. The purity of the sorted cell populations was not determined after isolation due to the limited numbers of cells recovered by FACS.
    Cd34, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 94/100, based on 10326 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    cd34  (Abcam)
    92
    Abcam cd34
    a The <t>CD34-positive</t> microvessels were represented by black arrows (original magnification × 100). b The number of CD34-positive vessels/mm 2 was 42.13 ± 2.59/mm 2 in the bradykinin group and 31.92 ± 1.40/mm 2 in the control group. n = 5 per group. * P
    Cd34, supplied by Abcam, used in various techniques. Bioz Stars score: 92/100, based on 2711 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Santa Cruz Biotechnology cd34
    The suppressive effects of miR-612 on HCC invasion, colonization, and distant metastasis. (A) Pathology analysis of tissue sections from recipient mice at week 6 after transplantation. (top) Hematoxylin and eosin (H E) staining; (bottom) anti–mouse <t>CD34</t> staining. Boxed area highlights intraliver dissemination and vascular invasion of tumor cells. (B and C) Lung colonization and outgrowth in recipient mice on day 42 after tail vein injection with 1.0 × 10 5 tumor cells. (B) Quantitation of tumor foci (IOD of fluorescent area) in lung of mice injected HCCLM3 and HepG2 cells. (C) Images showing the colonization and outgrowth foci of HCCLM3-RFP and HepG2-GFP cells in the lung of recipient mice. (D) Images showing the lung colonization in the recipient mice on day 3 after tail vein injection with 1.0 × 10 5 HCCLM3-RFP and HepG2-GFP cells. (E–G) Intraliver and lung metastasis in the recipient mice on day 42 after HCCLM3-RFP orthotopic transplantation. (E) Images showing metastatic foci in liver (top) and lung (bottom). (F) Quantitation of tumor foci (IOD of fluorescent area) in the liver and lung of recipient mice. (G) Incidence of intraliver dissemination and lung metastasis in WT, mock, and miR-612-o HCCLM3 xenografts. WT, mock, miR-612-o, or miR-612-i was defined as nontransfected, negative control oligonucleotide transfected, miR-612 mimic transfected, or miR-612 inhibitor transfected, respectively. Bars: (A) 100 µm; (C, fluorescent image) 2.0 mm; (C, H E image) 100 µm; (D) 100 µm; (E, liver) 6.0 mm; (E, lung) 2.0 mm. Data are mean ± SEM ( n = 5) and are representative of three independent experiments.
    Cd34, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 94/100, based on 2377 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Agilent technologies cd34
    Histology . Bland appearing spindle cells arranged in monotonous storiform pattern, infiltrating between lobules of fat in an honeycomb pattern in DFSP (H E; panel a). <t>CD34</t> (panel b) and Apolipoprotein-D (panel c) expression in DFSP. Malignat fibrous histicitoma-like areas in DFSP (DFSP-FS) (panel d). Loss of expression of CD 34 (panel e) and Apolopoprotein-D (panel f).
    Cd34, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 92/100, based on 2088 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Miltenyi Biotec cd34 microbead kit
    Capture of <t>CD34</t> + blast-derived exosomes directly from AML patients' plasma. A) Negatively stained electron microscope images of exosomes captured on CD34 <t>microbeads</t> (* shows a vacant <t>microbead).</t> B) Increasing AML plasma volumes were used for capture of CD34 + exosomes by microbeads and recovered exosomes were studied by Western blots. The graph shows a linear relationship between the input plasma volumes and pixel densities of captured and blotted CD34 + exosomes. C) Exosomes were captured from plasma samples obtained from five CD34 + AML patients and were analyzed by Western blots. The percentage of leukemic blasts in the peripheral circulation of each of the patients is shown. CD81 serves as the exosome marker. D) Removal of platelet-derived exosomes from plasma using anti-CD61 Ab-coated microbeads prior to capture of CD34 + exosomes. Exosomes captured with CD61 microbeads ( left : CD61 + ) and CD34 + exosomes captured after removing CD61 + exosomes ( right : CD61neg/CD34 + ) are shown in a representative Western blot of three evaluated.
    Cd34 Microbead Kit, supplied by Miltenyi Biotec, used in various techniques. Bioz Stars score: 99/100, based on 1293 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Becton Dickinson anti cd34
    ABT-737 has an equally high cytotoxic efficacy in disseminated AML pLSCs. Bone marrow and peripheral blood-derived primary AML cells from 2 matching patients were cultured and treated and the percentage of surviving cells determined as described before. (A) Viability of bone marrow-derived and peripheral blood-derived overall AML population after exposure to ABT-737. (B) The percentage of <t>CD34</t> + /CD38 − cells within the surviving population of leukemic blasts in both bone marrow and peripheral blood-derived matching sample sets showing equal efficacy of ABT-737 compound against AML blasts and pLSCs. The % of pLSCs within live AML cell population was evaluated as a number of CD34 + /CD38 − -7AAD − events. Different shapes indicate data-points corresponding to different patient samples. (Patients cross-referencing with Table 1 : Patient 2 (■), Patient 4 (□)).
    Anti Cd34, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 92/100, based on 1405 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Becton Dickinson cd34 pe
    Gating strategy for the enumeration of viable CD45 + [CD3/CD19/CD20/CRTH2] − CD16 − CD115 − CD11b + <t>CD34</t> + fibrocytes in the whole blood. Absolute counting beads were successfully resolved from cells and gated for counting. The representative assay was performed with the fresh blood sample of a patient with treatment-resistant asthma. 7-AAD, 7-amino-actinomycin-D; FSC, forward scatter; PB, Pacific Blue; SSC, side scatter.
    Cd34 Pe, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 94/100, based on 2141 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Becton Dickinson cd34 fitc
    Characterization of hMSCs. (A–C) Images of differentiated hMSCs after induction into specific tissues. (A) The lipid droplet accumulation in differentiated cells was visualized using Oil Red O staining after 2 weeks of adipogenic induction. (B) Calcium deposition was stained with Alizarin Red S after 2 weeks of osteogenic induction. (C) Glycosaminoglycans in cell pellets were revealed by Toluidine blue staining after 2 weeks of chondrogenic induction. (D) hMSCs (1×10 6 cells/ml) were stained with <t>FITC-</t> or PE- conjugated antibodies specific for human CD29, <t>CD34,</t> CD45, CD73, CD105, and HLA-DR.
    Cd34 Fitc, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 94/100, based on 1862 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Becton Dickinson cd34 apc
    Multicolour flow cytometric immunophenotypic analysis with monoclonal antibodies. Intraoral MSPCs at passages 3–7 were labelled with specific fluorochrome-conjugated monoclonal antibodies against the indicated surface antigens and analysed by flow cytometry. Analysed MSPCs demonstrated the same immunophenotype, with expression of (a) CD73, (b) CD90, and (c) CD105 but no expression of (d) CD14, (e) CD19, (f) <t>CD34,</t> (g) CD45, and (h) HLA-DR. A representative example of ten experiments is shown. PE: phycoerythrin; <t>APC:</t> allophycocyanin; PerCP: peridium-chlorophyll protein complex; FITC: fluorescein isothiocyanate.
    Cd34 Apc, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 92/100, based on 1230 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Novocastra cd34
    The tumor cells tested positive for alpha smooth muscle actin, calponin, H-caldesmon, but were negative for <t>CD34,</t> p53, estrogen receptor, progesterone receptor, and Epstein-Barr virus. In approximately 25% of all the tumor cells, the Ki67 proliferative index was positive. A: Pathological finding of thyroid gland stained (10 ×); B: Positive immunohistochemical staining for α-smooth muscle actin (10 ×); C: Negative immunohistochemical staining for P53 (20 ×); D: Pathological finding of 25% positive Ki67 index in tumor cells (20 ×); E: Negative immunohistochemical staining for CD34 (10 ×); F: Positive immunohistochemical staining for calponin (20 ×).
    Cd34, supplied by Novocastra, used in various techniques. Bioz Stars score: 92/100, based on 383 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher cd34 monoclonal antibody
    Lineage Tracing of Inter-Follicular Epidermis Stem Cells in Aspm-CreER Mice (A) Scheme of the epithelial skin structure with epidermis at the top and HF at the bottom. CL, cornified layer; G/SL, granular/spinous layer; BL, basal layer; SG, sebaceous gland; HFSC, HF stem cell; HG, hair germ; DP, dermal papilla. Note the marking of epidermis versus HF using Aspm -CreER versus Lgr5 -CreER genetic drivers. (B) Gene expression analysis of FACS-sorted inter-follicular epidermis (IFE) BL cells and HFSCs in Catagen (PD41) and Anagen (PD24). N = 3 of biological replicates; ∗ p = 0.009, ∗∗ p = 0.0002, ∗∗∗ p = 0.017, ∗∗∗∗ p = 0.001. HFSCs are FACS sorted as <t>α6-integrin+/CD34+</t> and BL cells were α6-integrin+/CD34− cells. Student’s t test was used for all significance tests. (C) Scheme of lineage tracing using Aspm -CreER;Rosa26-tdTomato mice after tamoxifen (TM) injection (red arrows) and different experimental endpoints (black arrowhead); 'sac' stands for time of sacrifice. (D) Back skin of Aspm -CreER;tdTomato mice treated as shown in (C) at endpoints indicated. Arrowhead shows Aspm -CreER-marked cells positive for tdTomato. Asterisk indicates autofluorescence from the hair shaft. N = 2 for TM−; N = 3 for the rest of the time points. (E) Experimental scheme and whole-mouse pictures showing back skin hair plucking of Aspm -CreER;tdTomato mice at time points indicated. Note that the right-hand side was plucked, and the hairs grew back by 2 weeks post-plucking. N = 3. (F) Lineage tracing of Aspm -CreER-marked positive IFE cells at PD98, 1 month after plucking. Left panel shows no migration in the HFs. Middle panel shows migration of Aspm cells to the infundibulum (arrowhead). Right panel shows migration of Aspm cells to the bulge (arrowhead). N = 3. The table was generated from averaging 100 HFs from 3 mice at 1 month after hair plucking. Scale bars, 20 μm.
    Cd34 Monoclonal Antibody, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 673 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abcam anti cd34
    In vivo inhibition of miR-138-5p reduces the pulmonary vascular cell proliferation in MCT-PH rats. a Immunofluorescence staining of frozen rat lung sections and confocal imaging with Click-iT 5-ethynyl-2′-deoxyuridine (EdU; white nuclei = EdU-positive nuclei = proliferating cells, yellow arrow) in combination with α-smooth muscle actin (α-SMA; in green, <t>CD34;</t> in red). Scale bar = 20 μm. The cells were counterstained with DAPI (blue). b Quantification of the percentage of proliferating cells in the lungs. ( n = 6 different rats per condition). * P
    Anti Cd34, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 508 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher anti cd34
    After gating for exclusion of debris and isolation of single cells, <t>CD45-/CD34+/CD31-</t> cells were isolated (21.9 ±± 2.4%). Flow cytometry gates were drawn based on unstained control ASCs.
    Anti Cd34, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 552 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Immunotec cd34
    Morphology and phenotype of CD40-activated <t>CD34</t> + HPC and in situ localization of CD1a − /CD40 − , DR + DC. CD34 + HPC prepared as described were seeded onto control (CD32L cells) cultures ( a ) or onto CD40L (CD40L-L cells) cultures ( b–i ) and evaluated at days 8 ( a and b ) and days 14 ( c – i ). Photomicrographs were taken from cells in culture wells ( a–d ) at ×200 ( a and b ) and ×400 ( c and d ) magnification, and from cytospin preparations ( e ) or from BioRad slides ( f–i ) at ×1,000 magnification. In ( c ) and ( d ), the same field was taken at a 15-s interval to show dendrite motility. ( e ) May– Grünwald staining of DC. ( f ) HIV–gp120 binding. ( g ) shows a CD26 + DC. ( h ) A rel-B + DC together with a rel-B − , nondendritic cell. ( i ) MHC class II DR + DC. ( j and k ) Tonsil sections stained for CD1a and CD40 (both in blue ) and MHC class II DR ( red ) showing single DR + , CD1a − and CD40 − DC scattered in the T cell area. ( j ) ×400 magnification and ( k ) ×1,000 magnification.
    Cd34, supplied by Immunotec, used in various techniques. Bioz Stars score: 93/100, based on 370 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Becton Dickinson anti cd34 pe
    Design and Validation of a Unified Vector Encoding iMC and PSCA.ζ CAR (A) Retroviral transgene design encoding MyD88, CD40, tandem FKBP12v36, T2A, signal peptide, A11 scFv variable light (V L ) and variable heavy (V H ) domains, the minimal <t>CD34</t> epitope, CD8α stalk and transmembrane region, and the CD3ζ signaling domain. (B) Flow cytometric analysis to determine transduction efficiency using anti-CD3 and anti-CD34 antibodies compared to non-transduced (NT) T cells. (C) iMC-PSCA.ζ-modified T cells only proliferate in the presence of exogenous IL-2 (100 U/mL) when stimulated with 10 nM rimiducid (Rim), but they demonstrate antigen-independent survival in the presence of IL-2 or with weekly iMC activation over 5 weeks (two individual donors shown). (D) T cells were transduced with EGFPluc or with both EGFPluc and iMC-PSCA.ζ-encoding-retrovirus and subsequently cultured with HPAC-RFP tumor cells at a 1:10 effector-to-tumor (E:T) cell ratio for 48 hr with variable Rim concentrations (0–250 nM) (n = 3). IL-6 and IL-2 production was measured by ELISA. (E) Control T cells (EGFPluc) or iMC-PSCA.ζ/EGFPluc-modified T cells were cultured at a 1:10 E:T ratio with HPAC-RFP cells at variable Rim concentrations (0–250 nM) for 7 days. Tumor cell killing (RFP; red) and T cell (EGFP; green) proliferation were measured by live-cell imaging using an IncuCyte imager. (F and G) EGFPluc and iMC-PSCA.ζ/EGFPluc-modified T cells were cultured with HPAC-RFP tumor cells at an E:T ratio of 1:20 and imaged over 7 days (n = 2). T cells transduced with iMC-PSCA.ζ/EGFPluc were stimulated with or without 2 nM Rim on day 0 of culture initiation.
    Anti Cd34 Pe, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 93/100, based on 636 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Ventana Medical cd34
    Immunohistochemical features of PLNTY. Strong labeling for GFAP as seen in case 3 ( a ), OLIG2 positivity as seen in case 2 ( b ), negativity for HuC/HuD as seen in case 6, and expression of BRAF V600E as seen in case 3. Diffuse strong expression of <t>CD34</t> in both tumor cells ( e , f ) and peripherally associated ramified neural elements ( g , h ) as seen in case 3 ( e , g ) and case 6 ( f , h )
    Cd34, supplied by Ventana Medical, used in various techniques. Bioz Stars score: 93/100, based on 271 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Pharmingen cd34
    Fibrocyte recruitment to kidney and spleen in response to ureteral obstruction. Male coll-GFP bone marrow was transplanted into female WT lethally irradiated mice. Chimerism was confirmed at 6 weeks by assessing leukocytes for Y chromosome. A: Image of perivascular GFP+ cells in d7 UUO kidney from coll-GFP ⇒WT bone marrow chimeric mice (magnification = original ×400). B: Confirmation of extensive fibrosis in this model by picrosirius red staining of d14 UUO kidney from chimeric mice. Note prominent perivascular fibrosis ( arrows ) as well as interstitial fibrosis. C–D: Image of d7 in UUO kidney from coll-GFP ⇒WT BM chimeric mice showing GFP+ cells co-expressing <t>CD34</t> and CD45, confirming these cells to be fibrocytes (magnification = original ×400). E–F: Image of d7 UUO kidney showing BM-derived coll1a1 -GFP+ cells lacking αSMA or S100A4 expression (magnification = original ×400). G: Red pulp of spleen from coll-GFP ⇒WT BM chimeric mice, 48 hours after UUO surgery, note the presence of GFP+ cells. H: Time course of fibrocyte recruitment to fibrotic kidney, spleen and full thickness skin wound. Number of cells is per sagittal section for kidney, transverse section of spleen, and transverse section of skin wound.
    Cd34, supplied by Pharmingen, used in various techniques. Bioz Stars score: 92/100, based on 385 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Becton Dickinson anti cd34 fitc
    Studying the effects of FGF2 on cellular calcium in satellite cells associated with host FDB fibers. Muscle cultures pre-stained with <t>anti-CD34-FITC</t> to identify satellite cells (A,C) were then loaded with calcium indicator X rhod-1 AM (E) . Transmitted light images are shown in (B,D) . Application of FGF2 at 2 ng/ml to the culture triggered a rise of calcium in the satellite cell in the confocal image plane ( F ; 9 satellite cells from 2 mice). Rinse with Ringer's solution without FGF2 reverses the rise in cell calcium ( G ; 9 cells from 2 mice). In cultures pre-loaded with calcium chelator BAPTA AM FGF2 caused little increase in calcium ( H ; 8 cells from 2 mice). In cultures rinsed with calcium free Ringer's solution, the effects of FGF2 are minimal ( I ; 8 cells from 2 mice). Incubation with TRPC blocker SKF 96365 attenuated the effects of FGF2 on cellular calcium ( J ; 8 satellite cells from 2 mice). FGF2 has no effects on cellular calcium of FDB fibers ( K ; 11 FDB fibers from 2 mice). ** p
    Anti Cd34 Fitc, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 91/100, based on 470 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Santa Cruz Biotechnology anti cd34
    Repressed HIF1A expression does not significantly affect tumor growth in SCLC cell xenografts U-1906 cells, transduced with shRNA against HIF1A (sh-H1) or a non-targeting control (sh-C), were injected subcutaneously into nude mice. ( A , B ) Mean tumor specimen weight and tumor volume, 14 days after injection. A, all tumors ( n = 12) and B, specimens with confirmed tumor growth ( n = 11, see text and Panels in E). Data are presented as mean ± SEM and statistical significance was determined using Student´s t -test. Data is from one of two independent experiments ( n = 6 for each group). ( C ) Sections of formalin-fixed paraffin-embedded xenografts were characterized immunohistochemically for expression of HIF-1α, Ki67 and <t>CD34.</t> ( D ) The corresponding dissected tumors are shown. ( E ) Example of a specimen, devoid of tumor cells, stained with hematoxylin-eosin (H/E) and anti-Ki67 antibody. Scale bars, 100 μm.
    Anti Cd34, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 92/100, based on 456 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Abcam anti cd34 antibody
    In vivo principle exploration of anti-tumor using this special treatment method. a LCSM images of PANC-1 tumor slices co-stained by TUNEL immunofluorescence <t>CD34</t> immunofluorescence that were harvested after sacrificing of PANC-1-bearing nude mice treated with aforementioned different treatments (G1-G5) on the 10th day post-treatment, scale bar: 50 μm, and TUNEL CD34 immunofluorescence co-staining was employed to determine cell apoptosis and vascular density, respectively, wherein FITC (green) and TRITC (red) were used to label TUNEL and CD34, respectively, and nuclei (blue) was stained by DAPI. b , c Semi-quantitative data of apoptotic cells ( b ) and apoptotic endothelial cells in all apoptotic cells ( c ) after different treatments with G1–G5 via calculating the ratios of green-labeled cells in whole blue-labeled cells and red and green co-labeled cells in all green-labeled cells according to the TUNEL immunofluorescence staining, respectively. Data are expressed as mean ± SD ( n = 3). * P
    Anti Cd34 Antibody, supplied by Abcam, used in various techniques. Bioz Stars score: 93/100, based on 275 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Agilent technologies anti cd34
    A representative case of conjunctival melanoma in a 65-year-old female. Notes: The pigmented tumor is noted in the corneal limbs with abnormal conjunctival vessels ( A ). Histopathology of the resected tumor reveals epithelioid-type melanoma cells infiltrated beneath the conjunctival epithelium ( B ). VEGF immunoreactivity is clearly detected in the cytoplasm of tumor cells as a greenish coloration ( C ). In contrast, <t>CD34-positive</t> blood vessels are not detected in the tumor tissue ( D ). A bar indicates 50 µm; magnification ×40.
    Anti Cd34, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 92/100, based on 375 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Significantly decreased colony-forming efficiency of Smad4 mutant keratinocytes. (A and B) Flow cytometry of α6-integrin and CD34 double-labeled keratinocytes from dorsal epidermis of wild-type and Smad4 mutant mice at P72. (C–J) Clonal

    Journal:

    Article Title: Disruption of Smad4 in Mouse Epidermis Leads to Depletion of Follicle Stem Cells

    doi: 10.1091/mbc.E08-07-0731

    Figure Lengend Snippet: Significantly decreased colony-forming efficiency of Smad4 mutant keratinocytes. (A and B) Flow cytometry of α6-integrin and CD34 double-labeled keratinocytes from dorsal epidermis of wild-type and Smad4 mutant mice at P72. (C–J) Clonal

    Article Snippet: Primary antibodies used in this study were K14 at 1:1000, K1 at 1:1000, involucrin at 1:1000 (Covance Laboratories, Madison, WI), Ki67 at 1:1000 (Abcam, Cambridge, MA), bromodeoxyuridine (BrdU) at 1:100 (Abcam), CD34 at 1:200 (eBioscience, San Diego, CA), K15 at 1:100 (Chemicon, Temecula, CA), active-β-catenin at 1:100 (Upstate Biotechnology, Lake Placid, NY), and p-Akt at 1:200 (Cell Signaling, Beverly, MA).

    Techniques: Mutagenesis, Flow Cytometry, Cytometry, Labeling, Mouse Assay

    Effect of variation in culture methods on phenotype. (A) Effect of FBS content the proportion of the human CDCs expressing markers of CPC (c-Kit), endothelial (CD31, CD34) and mesenchymal (CD90) origin. (B) Flow cytometry dot plot demonstrating the influence of endothelial media and standard cardiac explant media on the proportion of WK rat outgrowth cells expressing the CPC marker c-Kit.

    Journal: PLoS ONE

    Article Title: Validation of the Cardiosphere Method to Culture Cardiac Progenitor Cells from Myocardial Tissue

    doi: 10.1371/journal.pone.0007195

    Figure Lengend Snippet: Effect of variation in culture methods on phenotype. (A) Effect of FBS content the proportion of the human CDCs expressing markers of CPC (c-Kit), endothelial (CD31, CD34) and mesenchymal (CD90) origin. (B) Flow cytometry dot plot demonstrating the influence of endothelial media and standard cardiac explant media on the proportion of WK rat outgrowth cells expressing the CPC marker c-Kit.

    Article Snippet: The following monoclonal antibodies and conjugated fluorochromes were used with corresponding isotype controls: AA4 (AR32AA4, BD Pharmigen, San Jose, Ca), CD3 (MCA772FB, AbD Serotec, Oxford, UK), CD11b (MCA275FB, AbD Serotec), CD31 (sc-28188 Santa Cruz Biotech; BD Pharmingen 555445), CD34 (Chemicon CBL555F); CD45 (MCA340FB AbD Serotec; BD Pharmingen 555482), CD90-FITC (Dianova DIA120); c-Kit (sc-168 Santa Cruz Biotech; BD Pharmingen 550412), Ox-62 (MCA1029G, AbD Serotec).

    Techniques: Expressing, Flow Cytometry, Cytometry, Marker

    Flow cytometric analysis of the first outgrowth collection from cardiac tissue. (A) A comparison of the proportion of the initial human and WK rat outgrowth cells expressing markers of CPC (c-Kit), endothelial (CD31, CD34) and mesenchymal (CD90) origin. (B) Flow cytometry dot plot of the initial cellular outgrowth harvested from WK rat tissue demonstrating co-segregation of mast cell antigen AA4 with CPC marker c-Kit. (C) Flow cytometry dot plots of the initial outgrowth harvested from WK rat tissue demonstrating the co-segregation and abundance of c-Kit and CD45.

    Journal: PLoS ONE

    Article Title: Validation of the Cardiosphere Method to Culture Cardiac Progenitor Cells from Myocardial Tissue

    doi: 10.1371/journal.pone.0007195

    Figure Lengend Snippet: Flow cytometric analysis of the first outgrowth collection from cardiac tissue. (A) A comparison of the proportion of the initial human and WK rat outgrowth cells expressing markers of CPC (c-Kit), endothelial (CD31, CD34) and mesenchymal (CD90) origin. (B) Flow cytometry dot plot of the initial cellular outgrowth harvested from WK rat tissue demonstrating co-segregation of mast cell antigen AA4 with CPC marker c-Kit. (C) Flow cytometry dot plots of the initial outgrowth harvested from WK rat tissue demonstrating the co-segregation and abundance of c-Kit and CD45.

    Article Snippet: The following monoclonal antibodies and conjugated fluorochromes were used with corresponding isotype controls: AA4 (AR32AA4, BD Pharmigen, San Jose, Ca), CD3 (MCA772FB, AbD Serotec, Oxford, UK), CD11b (MCA275FB, AbD Serotec), CD31 (sc-28188 Santa Cruz Biotech; BD Pharmingen 555445), CD34 (Chemicon CBL555F); CD45 (MCA340FB AbD Serotec; BD Pharmingen 555482), CD90-FITC (Dianova DIA120); c-Kit (sc-168 Santa Cruz Biotech; BD Pharmingen 550412), Ox-62 (MCA1029G, AbD Serotec).

    Techniques: Flow Cytometry, Expressing, Cytometry, Marker

    FACS isolation of CD34 + cells infected with lentiviral vectors. Purified CD34 + cells (3 × 10 6 ) were infected with lentiviral vectors (MOI = 3) by centrifugation (4 h at 2,500 rpm) in a Beckman GPR centrifuge. Cells were incubated for 72 h in IMDM supplemented with 10% FBS and 30 μl of cytokine cocktail StemSpan CC100. Cells were incubated with a PE-conjugated human-specific MAb against the CD34 marker, and cell sorting was performed with a FACSVantage flow cytometer. An additional cell sample was incubated with a mouse immunoglobulin G γ isotype control antibody to set compensation and gates for FACS. The R2 gate in each panel represents the gate set to isolate CD34 + GFP + ). The percentage of CD34 + GFP + cells is displayed in the upper right panel. Mock-transduced CD34 + cells were isolated only on the basis of CD34 + expression and are represented by the R3 gate. LVs used in each transduction are as follows: GFP, HR′CMV-GFP; Tax1, HR′CMV-Tax1/GFP; Tax1(−), HR′CMV-Tax1(−)/GFP; Tax2, HR′CMV-Tax2/GFP; and Vpr, HR′CMV-Vpr/GFP. The purity of the sorted cell populations was not determined after isolation due to the limited numbers of cells recovered by FACS.

    Journal: Journal of Virology

    Article Title: Human T-Cell Leukemia Virus Type 1 Tax Oncoprotein Suppression of Multilineage Hematopoiesis of CD34+ Cells In Vitro

    doi: 10.1128/JVI.77.22.12152-12164.2003

    Figure Lengend Snippet: FACS isolation of CD34 + cells infected with lentiviral vectors. Purified CD34 + cells (3 × 10 6 ) were infected with lentiviral vectors (MOI = 3) by centrifugation (4 h at 2,500 rpm) in a Beckman GPR centrifuge. Cells were incubated for 72 h in IMDM supplemented with 10% FBS and 30 μl of cytokine cocktail StemSpan CC100. Cells were incubated with a PE-conjugated human-specific MAb against the CD34 marker, and cell sorting was performed with a FACSVantage flow cytometer. An additional cell sample was incubated with a mouse immunoglobulin G γ isotype control antibody to set compensation and gates for FACS. The R2 gate in each panel represents the gate set to isolate CD34 + GFP + ). The percentage of CD34 + GFP + cells is displayed in the upper right panel. Mock-transduced CD34 + cells were isolated only on the basis of CD34 + expression and are represented by the R3 gate. LVs used in each transduction are as follows: GFP, HR′CMV-GFP; Tax1, HR′CMV-Tax1/GFP; Tax1(−), HR′CMV-Tax1(−)/GFP; Tax2, HR′CMV-Tax2/GFP; and Vpr, HR′CMV-Vpr/GFP. The purity of the sorted cell populations was not determined after isolation due to the limited numbers of cells recovered by FACS.

    Article Snippet: CD34+ cells transduced with Tax1 demonstrated a two- to fivefold reduction in clonogenic colony-forming activity in vitro, in comparison with CD34+ cells transduced with LVs encoding Tax2, Tax1(−), or only GFP (Fig. ).

    Techniques: FACS, Isolation, Infection, Purification, Centrifugation, Incubation, Marker, Flow Cytometry, Cytometry, Expressing, Transduction

    Clonogenic colony-forming activity of lentiviral vector- transduced CD34 + cells. GFP + CD34 + cells were purified by FACS, and isolated cells (10 3 ). (A) Total number of myeloid, erythroid and pluripotential colonies per CD34 + GFP + cells (10 3 ) plated was determined at 14 days postplating. Purified CD34 + GFP + cell samples were plated in triplicate. Each column represents a separate sorting experiment. Colony-forming activities were assayed four times for each transduction, except for Tax1(−) and Tax2-transduced CD34 + cells, which were assayed three times (sorting experiments 2, 3, and 4). Transduction of CD34 + cells with HR′CMV-Vpr/GFP resulted in no colony formation in FACS experiments 1 and 4 and is indicated by an asterisk. (B) Relative distribution of clonogenic colonies. Colonies were analyzed by morphology and characterized as CFU-GM, BFU-E, or HPP-CFC. The average numbers of CFU-GM colonies that arose per 10 3 purified CD34 + GFP + cells plated were 38.8 (Mock), 24.4 (GFP), 7.3 (Tax1), 21.2 [Tax1(−)], and 21.4 (Tax2). The average numbers of BFU-E colonies arising per 10 3 CD34 + GFP+ cells plated were 14.9 (Mock), 9.0 (GFP), 2.8 (Tax1), 8.0 [Tax1(−)], and 8.3 (Tax2). The average numbers of CFU-HPP colonies arising per 10 3 CD34 + GFP + cells plated were 6.0 (Mock), 3.6 (GFP), 1.2 (Tax1), 3.2 [Tax1(−)], and 3.3 (Tax2). Statistical analysis was performed by ANOVA ( P

    Journal: Journal of Virology

    Article Title: Human T-Cell Leukemia Virus Type 1 Tax Oncoprotein Suppression of Multilineage Hematopoiesis of CD34+ Cells In Vitro

    doi: 10.1128/JVI.77.22.12152-12164.2003

    Figure Lengend Snippet: Clonogenic colony-forming activity of lentiviral vector- transduced CD34 + cells. GFP + CD34 + cells were purified by FACS, and isolated cells (10 3 ). (A) Total number of myeloid, erythroid and pluripotential colonies per CD34 + GFP + cells (10 3 ) plated was determined at 14 days postplating. Purified CD34 + GFP + cell samples were plated in triplicate. Each column represents a separate sorting experiment. Colony-forming activities were assayed four times for each transduction, except for Tax1(−) and Tax2-transduced CD34 + cells, which were assayed three times (sorting experiments 2, 3, and 4). Transduction of CD34 + cells with HR′CMV-Vpr/GFP resulted in no colony formation in FACS experiments 1 and 4 and is indicated by an asterisk. (B) Relative distribution of clonogenic colonies. Colonies were analyzed by morphology and characterized as CFU-GM, BFU-E, or HPP-CFC. The average numbers of CFU-GM colonies that arose per 10 3 purified CD34 + GFP + cells plated were 38.8 (Mock), 24.4 (GFP), 7.3 (Tax1), 21.2 [Tax1(−)], and 21.4 (Tax2). The average numbers of BFU-E colonies arising per 10 3 CD34 + GFP+ cells plated were 14.9 (Mock), 9.0 (GFP), 2.8 (Tax1), 8.0 [Tax1(−)], and 8.3 (Tax2). The average numbers of CFU-HPP colonies arising per 10 3 CD34 + GFP + cells plated were 6.0 (Mock), 3.6 (GFP), 1.2 (Tax1), 3.2 [Tax1(−)], and 3.3 (Tax2). Statistical analysis was performed by ANOVA ( P

    Article Snippet: CD34+ cells transduced with Tax1 demonstrated a two- to fivefold reduction in clonogenic colony-forming activity in vitro, in comparison with CD34+ cells transduced with LVs encoding Tax2, Tax1(−), or only GFP (Fig. ).

    Techniques: Activity Assay, Plasmid Preparation, Purification, FACS, Isolation, Transduction

    a The CD34-positive microvessels were represented by black arrows (original magnification × 100). b The number of CD34-positive vessels/mm 2 was 42.13 ± 2.59/mm 2 in the bradykinin group and 31.92 ± 1.40/mm 2 in the control group. n = 5 per group. * P

    Journal: World Journal of Surgical Oncology

    Article Title: Effects of bradykinin on the survival of multiterritory perforator flaps in rats

    doi: 10.1186/s12957-019-1570-3

    Figure Lengend Snippet: a The CD34-positive microvessels were represented by black arrows (original magnification × 100). b The number of CD34-positive vessels/mm 2 was 42.13 ± 2.59/mm 2 in the bradykinin group and 31.92 ± 1.40/mm 2 in the control group. n = 5 per group. * P

    Article Snippet: Endothelial cells can be labeled by CD34.

    Techniques:

    The suppressive effects of miR-612 on HCC invasion, colonization, and distant metastasis. (A) Pathology analysis of tissue sections from recipient mice at week 6 after transplantation. (top) Hematoxylin and eosin (H E) staining; (bottom) anti–mouse CD34 staining. Boxed area highlights intraliver dissemination and vascular invasion of tumor cells. (B and C) Lung colonization and outgrowth in recipient mice on day 42 after tail vein injection with 1.0 × 10 5 tumor cells. (B) Quantitation of tumor foci (IOD of fluorescent area) in lung of mice injected HCCLM3 and HepG2 cells. (C) Images showing the colonization and outgrowth foci of HCCLM3-RFP and HepG2-GFP cells in the lung of recipient mice. (D) Images showing the lung colonization in the recipient mice on day 3 after tail vein injection with 1.0 × 10 5 HCCLM3-RFP and HepG2-GFP cells. (E–G) Intraliver and lung metastasis in the recipient mice on day 42 after HCCLM3-RFP orthotopic transplantation. (E) Images showing metastatic foci in liver (top) and lung (bottom). (F) Quantitation of tumor foci (IOD of fluorescent area) in the liver and lung of recipient mice. (G) Incidence of intraliver dissemination and lung metastasis in WT, mock, and miR-612-o HCCLM3 xenografts. WT, mock, miR-612-o, or miR-612-i was defined as nontransfected, negative control oligonucleotide transfected, miR-612 mimic transfected, or miR-612 inhibitor transfected, respectively. Bars: (A) 100 µm; (C, fluorescent image) 2.0 mm; (C, H E image) 100 µm; (D) 100 µm; (E, liver) 6.0 mm; (E, lung) 2.0 mm. Data are mean ± SEM ( n = 5) and are representative of three independent experiments.

    Journal: The Journal of Experimental Medicine

    Article Title: miR-612 suppresses the invasive-metastatic cascade in hepatocellular carcinoma

    doi: 10.1084/jem.20120153

    Figure Lengend Snippet: The suppressive effects of miR-612 on HCC invasion, colonization, and distant metastasis. (A) Pathology analysis of tissue sections from recipient mice at week 6 after transplantation. (top) Hematoxylin and eosin (H E) staining; (bottom) anti–mouse CD34 staining. Boxed area highlights intraliver dissemination and vascular invasion of tumor cells. (B and C) Lung colonization and outgrowth in recipient mice on day 42 after tail vein injection with 1.0 × 10 5 tumor cells. (B) Quantitation of tumor foci (IOD of fluorescent area) in lung of mice injected HCCLM3 and HepG2 cells. (C) Images showing the colonization and outgrowth foci of HCCLM3-RFP and HepG2-GFP cells in the lung of recipient mice. (D) Images showing the lung colonization in the recipient mice on day 3 after tail vein injection with 1.0 × 10 5 HCCLM3-RFP and HepG2-GFP cells. (E–G) Intraliver and lung metastasis in the recipient mice on day 42 after HCCLM3-RFP orthotopic transplantation. (E) Images showing metastatic foci in liver (top) and lung (bottom). (F) Quantitation of tumor foci (IOD of fluorescent area) in the liver and lung of recipient mice. (G) Incidence of intraliver dissemination and lung metastasis in WT, mock, and miR-612-o HCCLM3 xenografts. WT, mock, miR-612-o, or miR-612-i was defined as nontransfected, negative control oligonucleotide transfected, miR-612 mimic transfected, or miR-612 inhibitor transfected, respectively. Bars: (A) 100 µm; (C, fluorescent image) 2.0 mm; (C, H E image) 100 µm; (D) 100 µm; (E, liver) 6.0 mm; (E, lung) 2.0 mm. Data are mean ± SEM ( n = 5) and are representative of three independent experiments.

    Article Snippet: Density of target proteins, except CD34 used for vascular labeling, was determined as previously reported ( ).

    Techniques: Mouse Assay, Transplantation Assay, Staining, Injection, Quantitation Assay, Negative Control, Transfection

    Histology . Bland appearing spindle cells arranged in monotonous storiform pattern, infiltrating between lobules of fat in an honeycomb pattern in DFSP (H E; panel a). CD34 (panel b) and Apolipoprotein-D (panel c) expression in DFSP. Malignat fibrous histicitoma-like areas in DFSP (DFSP-FS) (panel d). Loss of expression of CD 34 (panel e) and Apolopoprotein-D (panel f).

    Journal: Clinical Sarcoma Research

    Article Title: Fibrosarcomatous changes and expression of CD34+ and apolipoprotein-D in dermatofibrosarcoma protuberans

    doi: 10.1186/2045-3329-2-4

    Figure Lengend Snippet: Histology . Bland appearing spindle cells arranged in monotonous storiform pattern, infiltrating between lobules of fat in an honeycomb pattern in DFSP (H E; panel a). CD34 (panel b) and Apolipoprotein-D (panel c) expression in DFSP. Malignat fibrous histicitoma-like areas in DFSP (DFSP-FS) (panel d). Loss of expression of CD 34 (panel e) and Apolopoprotein-D (panel f).

    Article Snippet: Immunohistochemical expression of CD34 (Qbend-10, 1-100 dilution, Dako, Carpinteria CA, USA) and Apo-D (36C6, 1-200 dilution, Novocastra, New-castle-on-Tyne, UK) were assessed by two pathologists (M-G. and L.Z.) in all patients with adequate tumor tissue.

    Techniques: Expressing

    Five-year event free survival (EFS) according to CD34 and Apolipoprotein-D expression .

    Journal: Clinical Sarcoma Research

    Article Title: Fibrosarcomatous changes and expression of CD34+ and apolipoprotein-D in dermatofibrosarcoma protuberans

    doi: 10.1186/2045-3329-2-4

    Figure Lengend Snippet: Five-year event free survival (EFS) according to CD34 and Apolipoprotein-D expression .

    Article Snippet: Immunohistochemical expression of CD34 (Qbend-10, 1-100 dilution, Dako, Carpinteria CA, USA) and Apo-D (36C6, 1-200 dilution, Novocastra, New-castle-on-Tyne, UK) were assessed by two pathologists (M-G. and L.Z.) in all patients with adequate tumor tissue.

    Techniques: Expressing

    Capture of CD34 + blast-derived exosomes directly from AML patients' plasma. A) Negatively stained electron microscope images of exosomes captured on CD34 microbeads (* shows a vacant microbead). B) Increasing AML plasma volumes were used for capture of CD34 + exosomes by microbeads and recovered exosomes were studied by Western blots. The graph shows a linear relationship between the input plasma volumes and pixel densities of captured and blotted CD34 + exosomes. C) Exosomes were captured from plasma samples obtained from five CD34 + AML patients and were analyzed by Western blots. The percentage of leukemic blasts in the peripheral circulation of each of the patients is shown. CD81 serves as the exosome marker. D) Removal of platelet-derived exosomes from plasma using anti-CD61 Ab-coated microbeads prior to capture of CD34 + exosomes. Exosomes captured with CD61 microbeads ( left : CD61 + ) and CD34 + exosomes captured after removing CD61 + exosomes ( right : CD61neg/CD34 + ) are shown in a representative Western blot of three evaluated.

    Journal: PLoS ONE

    Article Title: Isolation and Characterization of CD34+ Blast-Derived Exosomes in Acute Myeloid Leukemia

    doi: 10.1371/journal.pone.0103310

    Figure Lengend Snippet: Capture of CD34 + blast-derived exosomes directly from AML patients' plasma. A) Negatively stained electron microscope images of exosomes captured on CD34 microbeads (* shows a vacant microbead). B) Increasing AML plasma volumes were used for capture of CD34 + exosomes by microbeads and recovered exosomes were studied by Western blots. The graph shows a linear relationship between the input plasma volumes and pixel densities of captured and blotted CD34 + exosomes. C) Exosomes were captured from plasma samples obtained from five CD34 + AML patients and were analyzed by Western blots. The percentage of leukemic blasts in the peripheral circulation of each of the patients is shown. CD81 serves as the exosome marker. D) Removal of platelet-derived exosomes from plasma using anti-CD61 Ab-coated microbeads prior to capture of CD34 + exosomes. Exosomes captured with CD61 microbeads ( left : CD61 + ) and CD34 + exosomes captured after removing CD61 + exosomes ( right : CD61neg/CD34 + ) are shown in a representative Western blot of three evaluated.

    Article Snippet: Immunoaffinity capture of exosomes from plasma To isolate blast-derived exosomes from plasma of patients with CD34+ blasts, the CD34 microbead kit (Miltenyi Biotec, Auburn, CA, USA) was used.

    Techniques: Derivative Assay, Staining, Microscopy, Western Blot, Marker

    Calibration experiments with CD34 + Kasumi-1 exosomes isolated by ultracentrifugation. A) Isolated exosomes were loaded at increasing protein concentrations onto gels and blotted using anti-CD34 Ab. Intensity of each band was measured in pixels. The graph illustrates a linear relationship between exosomal protein levels and pixel intensity in Western blots. B) Isolated exosomes (20 µg protein) were added to increasing volumes of CD34 microbeads (left ). Five or 10 µL of microbeads were sufficient to capture 20 µg of exosomes. Next, a 10 µL aliquot of beads was used to capture increasing concentrations of Kasumi-1 exosomes ( right ). The graph shows that the capacity of CD34 microbeads to capture up to 80 µg of input exosomes increased linearly. However, at 80 µg only about 40% of exosomal proteins were captured, suggesting that additional microbeads are necessary for capture of all Kasumi-1 exosomes.

    Journal: PLoS ONE

    Article Title: Isolation and Characterization of CD34+ Blast-Derived Exosomes in Acute Myeloid Leukemia

    doi: 10.1371/journal.pone.0103310

    Figure Lengend Snippet: Calibration experiments with CD34 + Kasumi-1 exosomes isolated by ultracentrifugation. A) Isolated exosomes were loaded at increasing protein concentrations onto gels and blotted using anti-CD34 Ab. Intensity of each band was measured in pixels. The graph illustrates a linear relationship between exosomal protein levels and pixel intensity in Western blots. B) Isolated exosomes (20 µg protein) were added to increasing volumes of CD34 microbeads (left ). Five or 10 µL of microbeads were sufficient to capture 20 µg of exosomes. Next, a 10 µL aliquot of beads was used to capture increasing concentrations of Kasumi-1 exosomes ( right ). The graph shows that the capacity of CD34 microbeads to capture up to 80 µg of input exosomes increased linearly. However, at 80 µg only about 40% of exosomal proteins were captured, suggesting that additional microbeads are necessary for capture of all Kasumi-1 exosomes.

    Article Snippet: Immunoaffinity capture of exosomes from plasma To isolate blast-derived exosomes from plasma of patients with CD34+ blasts, the CD34 microbead kit (Miltenyi Biotec, Auburn, CA, USA) was used.

    Techniques: Isolation, Western Blot

    The strategy for capture of CD34 + exosomes using anti-CD34 Ab-coated microbeads purchased from Miltenyi (A) and a schema for capture of CD34 + exosomes directly from AML patients' plasma (B).

    Journal: PLoS ONE

    Article Title: Isolation and Characterization of CD34+ Blast-Derived Exosomes in Acute Myeloid Leukemia

    doi: 10.1371/journal.pone.0103310

    Figure Lengend Snippet: The strategy for capture of CD34 + exosomes using anti-CD34 Ab-coated microbeads purchased from Miltenyi (A) and a schema for capture of CD34 + exosomes directly from AML patients' plasma (B).

    Article Snippet: Immunoaffinity capture of exosomes from plasma To isolate blast-derived exosomes from plasma of patients with CD34+ blasts, the CD34 microbead kit (Miltenyi Biotec, Auburn, CA, USA) was used.

    Techniques:

    Capture of CD34+ exosomes from total exosomal fractions by CD34+ microbeads. Isolated Kasumi-1 exosomes (Kas) and total exosome fractions isolated by ultracentrifugation from normal donor plasma (NC) or from CD34 + AML patient plasma (AML) were used for capture with CD34 microbeads. After the 1 st capture and removal of beads, 2 nd capture was performed with a fresh aliquot of CD34 microbeads. The final unbound fractions were ultracentrifuged to collect remaining exosomes. While the 2 nd capture was necessary to recover all CD34 + Kasumi-1 exosomes, a single capture was sufficient to recover all CD34 + exosomes from the bulk of exosomes isolated from the AML plasma. There were no CD34 + exosomes captured from the bulk exosomal fraction isolated from a normal donor's plasma. CD81 expression indicates that the unbound fractions still contain CD34 neg exosomes.

    Journal: PLoS ONE

    Article Title: Isolation and Characterization of CD34+ Blast-Derived Exosomes in Acute Myeloid Leukemia

    doi: 10.1371/journal.pone.0103310

    Figure Lengend Snippet: Capture of CD34+ exosomes from total exosomal fractions by CD34+ microbeads. Isolated Kasumi-1 exosomes (Kas) and total exosome fractions isolated by ultracentrifugation from normal donor plasma (NC) or from CD34 + AML patient plasma (AML) were used for capture with CD34 microbeads. After the 1 st capture and removal of beads, 2 nd capture was performed with a fresh aliquot of CD34 microbeads. The final unbound fractions were ultracentrifuged to collect remaining exosomes. While the 2 nd capture was necessary to recover all CD34 + Kasumi-1 exosomes, a single capture was sufficient to recover all CD34 + exosomes from the bulk of exosomes isolated from the AML plasma. There were no CD34 + exosomes captured from the bulk exosomal fraction isolated from a normal donor's plasma. CD81 expression indicates that the unbound fractions still contain CD34 neg exosomes.

    Article Snippet: Immunoaffinity capture of exosomes from plasma To isolate blast-derived exosomes from plasma of patients with CD34+ blasts, the CD34 microbead kit (Miltenyi Biotec, Auburn, CA, USA) was used.

    Techniques: Isolation, Expressing

    Schematic representation of the two-step immunomagnetic microbead-based cell separation protocol. Total dermal cells obtained from the processing of healthy human skin samples were subjected to magnetic-activated cell sorting separation. The first step was carried out in order to isolate CD31+ cells (i.e., putative endothelial cells), while the second step, performed on the pool of the remaining CD31− cells, allowed the further separation of CD31−/CD34+ cells (i.e., putative telocytes) from CD31−/CD34− cells (i.e., putative fibroblasts).

    Journal: International Journal of Molecular Sciences

    Article Title: A Two-Step Immunomagnetic Microbead-Based Method for the Isolation of Human Primary Skin Telocytes/CD34+ Stromal Cells

    doi: 10.3390/ijms21165877

    Figure Lengend Snippet: Schematic representation of the two-step immunomagnetic microbead-based cell separation protocol. Total dermal cells obtained from the processing of healthy human skin samples were subjected to magnetic-activated cell sorting separation. The first step was carried out in order to isolate CD31+ cells (i.e., putative endothelial cells), while the second step, performed on the pool of the remaining CD31− cells, allowed the further separation of CD31−/CD34+ cells (i.e., putative telocytes) from CD31−/CD34− cells (i.e., putative fibroblasts).

    Article Snippet: CD31−/CD34+ Telocyte Isolation CD31−/CD34+ TCs were isolated using the CD34 MicroBead Kit (catalog no. 130-046-702; Miltenyi Biotec, Bergisch Gladbach, Germany).

    Techniques: FACS

    Characterization of extracellular vesicles (EV) released from mesenchymal stromal cells (MSC). Size distribution (nm) and concentration (particles × 10 10 per milliliter) quantified by nanoparticle tracking analysis. (A): Representative graph for one sample of EV obtained from 3 × 10 6 MSC. (B): EV samples contrasted with uranyl‐oxalate solution and examined under a transmission electron microscope. Scale bar = 200 nm, original magnification ×8,000. Representative image of flow cytometry characterization of EV in one sample. Gates of EV were defined in the forward and side scatter dot plot by setting the upper size limit in 1 μm. Microbeads with a diameter of 1 μm were used for that purpose. The other dot plots represent EV stained with hematopoietic markers (CD34 and CD45), mesenchymal markers (CD90 and CD44), and exosomes markers (CD81 and CD63). Unstained controls are shown in gray, and EV stained with different antibodies are represented in black. (C): Samples were acquired on a FACS Calibur flow cytometer. (D): EV characterization by Western blot assay for the expression of CD63. Abbreviations: SSC, side scatter; FSC, forward scatter.

    Journal: Stem Cells (Dayton, Ohio)

    Article Title: The Incorporation of Extracellular Vesicles from Mesenchymal Stromal Cells Into CD34+ Cells Increases Their Clonogenic Capacity and Bone Marrow Lodging Ability

    doi: 10.1002/stem.3032

    Figure Lengend Snippet: Characterization of extracellular vesicles (EV) released from mesenchymal stromal cells (MSC). Size distribution (nm) and concentration (particles × 10 10 per milliliter) quantified by nanoparticle tracking analysis. (A): Representative graph for one sample of EV obtained from 3 × 10 6 MSC. (B): EV samples contrasted with uranyl‐oxalate solution and examined under a transmission electron microscope. Scale bar = 200 nm, original magnification ×8,000. Representative image of flow cytometry characterization of EV in one sample. Gates of EV were defined in the forward and side scatter dot plot by setting the upper size limit in 1 μm. Microbeads with a diameter of 1 μm were used for that purpose. The other dot plots represent EV stained with hematopoietic markers (CD34 and CD45), mesenchymal markers (CD90 and CD44), and exosomes markers (CD81 and CD63). Unstained controls are shown in gray, and EV stained with different antibodies are represented in black. (C): Samples were acquired on a FACS Calibur flow cytometer. (D): EV characterization by Western blot assay for the expression of CD63. Abbreviations: SSC, side scatter; FSC, forward scatter.

    Article Snippet: CD34+ cells were purified by immunomagnetic sorting in an AutoMACS (MiltenyiBiotec GmbH) after labeling with the human CD34 MicroBead Kit (MiltenyiBiotec).

    Techniques: Concentration Assay, Transmission Assay, Microscopy, Flow Cytometry, Cytometry, Staining, FACS, Western Blot, Expressing

    ABT-737 has an equally high cytotoxic efficacy in disseminated AML pLSCs. Bone marrow and peripheral blood-derived primary AML cells from 2 matching patients were cultured and treated and the percentage of surviving cells determined as described before. (A) Viability of bone marrow-derived and peripheral blood-derived overall AML population after exposure to ABT-737. (B) The percentage of CD34 + /CD38 − cells within the surviving population of leukemic blasts in both bone marrow and peripheral blood-derived matching sample sets showing equal efficacy of ABT-737 compound against AML blasts and pLSCs. The % of pLSCs within live AML cell population was evaluated as a number of CD34 + /CD38 − -7AAD − events. Different shapes indicate data-points corresponding to different patient samples. (Patients cross-referencing with Table 1 : Patient 2 (■), Patient 4 (□)).

    Journal: Leukemia Research Reports

    Article Title: The BH3-mimetic ABT-737 effectively kills acute myeloid leukemia initiating cells

    doi: 10.1016/j.lrr.2014.06.001

    Figure Lengend Snippet: ABT-737 has an equally high cytotoxic efficacy in disseminated AML pLSCs. Bone marrow and peripheral blood-derived primary AML cells from 2 matching patients were cultured and treated and the percentage of surviving cells determined as described before. (A) Viability of bone marrow-derived and peripheral blood-derived overall AML population after exposure to ABT-737. (B) The percentage of CD34 + /CD38 − cells within the surviving population of leukemic blasts in both bone marrow and peripheral blood-derived matching sample sets showing equal efficacy of ABT-737 compound against AML blasts and pLSCs. The % of pLSCs within live AML cell population was evaluated as a number of CD34 + /CD38 − -7AAD − events. Different shapes indicate data-points corresponding to different patient samples. (Patients cross-referencing with Table 1 : Patient 2 (■), Patient 4 (□)).

    Article Snippet: 2.4 Flow cytometry MNCs were incubated with anti-CD34 and anti-CD38 antibodies (BD Bioscience, San Diego, USA) in PBS/1% BSA (Sigma) following the manufacturer׳s instructions.

    Techniques: Derivative Assay, Cell Culture

    AML cells show variable sensitivity to the combination of cytarabine and daunorubicin. Bone marrow primary AML cells were cultured on a stromal-feeder layer and then were treated with 4.5 μM cytarabine (AraC)+1.35 μM daunorubicin (DnR) for 24 h. AML cell viability was evaluated using 7-AAD staining and flow cytometry. (A) The cytotoxic effect of AraC+DnR on the overall blast population. The graph shows the percentage of live cells for each sample. (B) Treatment with AraC+DnR leads to enrichment of the CD34 + /CD38 − cell population in the surviving fraction. The graph shows the percentage of the CD34 + /CD38 − cells within the surviving AML cell population in each sample. Different shapes indicate data-points corresponding to different patient samples. (Patients cross-referencing with Table 1 : Patient 1 (○), Patient 2 (■), Patient 3 (●), Patient 4 (□)).

    Journal: Leukemia Research Reports

    Article Title: The BH3-mimetic ABT-737 effectively kills acute myeloid leukemia initiating cells

    doi: 10.1016/j.lrr.2014.06.001

    Figure Lengend Snippet: AML cells show variable sensitivity to the combination of cytarabine and daunorubicin. Bone marrow primary AML cells were cultured on a stromal-feeder layer and then were treated with 4.5 μM cytarabine (AraC)+1.35 μM daunorubicin (DnR) for 24 h. AML cell viability was evaluated using 7-AAD staining and flow cytometry. (A) The cytotoxic effect of AraC+DnR on the overall blast population. The graph shows the percentage of live cells for each sample. (B) Treatment with AraC+DnR leads to enrichment of the CD34 + /CD38 − cell population in the surviving fraction. The graph shows the percentage of the CD34 + /CD38 − cells within the surviving AML cell population in each sample. Different shapes indicate data-points corresponding to different patient samples. (Patients cross-referencing with Table 1 : Patient 1 (○), Patient 2 (■), Patient 3 (●), Patient 4 (□)).

    Article Snippet: 2.4 Flow cytometry MNCs were incubated with anti-CD34 and anti-CD38 antibodies (BD Bioscience, San Diego, USA) in PBS/1% BSA (Sigma) following the manufacturer׳s instructions.

    Techniques: Cell Culture, Staining, Flow Cytometry, Cytometry

    ABT-737 has a potent cytotoxic effect on bone marrow-derived AML pLSCs. Bone marrow-derived primary AML cells were cultured on a stromal-feeder layer for 24 h and then treated with 30 nM and 300 nM of ABT-737 for 24 h. AML cell viability was evaluated using7-AAD staining and flow cytometry. (A) The cytotoxic effect of ABT-737 on the overall blast population. The graph shows the percentage of live cells for each sample. (B) ABT-737 is equally efficient in eradicating AML pLSCs. The graph shows the percentage of the CD34 + /CD38 − cells within the surviving AML cell population in each sample. E Horizontal lines indicate mean values. Different shapes indicate data-points corresponding to particular patient samples (Patients cross-referencing with Table 1 : Patient 1 (○), Patient 2 (■), Patient 3 (•), Patient 4 (□)).

    Journal: Leukemia Research Reports

    Article Title: The BH3-mimetic ABT-737 effectively kills acute myeloid leukemia initiating cells

    doi: 10.1016/j.lrr.2014.06.001

    Figure Lengend Snippet: ABT-737 has a potent cytotoxic effect on bone marrow-derived AML pLSCs. Bone marrow-derived primary AML cells were cultured on a stromal-feeder layer for 24 h and then treated with 30 nM and 300 nM of ABT-737 for 24 h. AML cell viability was evaluated using7-AAD staining and flow cytometry. (A) The cytotoxic effect of ABT-737 on the overall blast population. The graph shows the percentage of live cells for each sample. (B) ABT-737 is equally efficient in eradicating AML pLSCs. The graph shows the percentage of the CD34 + /CD38 − cells within the surviving AML cell population in each sample. E Horizontal lines indicate mean values. Different shapes indicate data-points corresponding to particular patient samples (Patients cross-referencing with Table 1 : Patient 1 (○), Patient 2 (■), Patient 3 (•), Patient 4 (□)).

    Article Snippet: 2.4 Flow cytometry MNCs were incubated with anti-CD34 and anti-CD38 antibodies (BD Bioscience, San Diego, USA) in PBS/1% BSA (Sigma) following the manufacturer׳s instructions.

    Techniques: Derivative Assay, Cell Culture, Staining, Flow Cytometry, Cytometry

    Gating strategy for the enumeration of viable CD45 + [CD3/CD19/CD20/CRTH2] − CD16 − CD115 − CD11b + CD34 + fibrocytes in the whole blood. Absolute counting beads were successfully resolved from cells and gated for counting. The representative assay was performed with the fresh blood sample of a patient with treatment-resistant asthma. 7-AAD, 7-amino-actinomycin-D; FSC, forward scatter; PB, Pacific Blue; SSC, side scatter.

    Journal: BBA Clinical

    Article Title: Enumeration of circulating fibrocytes for clinical use in asthma by an optimized single-platform flow cytometry assay

    doi: 10.1016/j.bbacli.2014.06.002

    Figure Lengend Snippet: Gating strategy for the enumeration of viable CD45 + [CD3/CD19/CD20/CRTH2] − CD16 − CD115 − CD11b + CD34 + fibrocytes in the whole blood. Absolute counting beads were successfully resolved from cells and gated for counting. The representative assay was performed with the fresh blood sample of a patient with treatment-resistant asthma. 7-AAD, 7-amino-actinomycin-D; FSC, forward scatter; PB, Pacific Blue; SSC, side scatter.

    Article Snippet: One hundred μl of treated or untreated blood were added to a 5-ml round-bottom tube containing 2 μl of the viability dye 7-amino-actinomycin-D (7-AAD, BD Biosciences, San Jose, CA, USA), which is detected on the PerCP-Cy5.5 channel, and appropriate dilutions in stain buffer (BD Biosciences) of the following fluorochrome-labeled monoclonal antibodies (all mouse IgG1κ ): 2.5 μl CD3-PerCP-Cy5.5, 2.5 μl CD19-PerCP-Cy5.5, 2.5 μl CD20-PerCP-Cy5.5, 2.5 μl CRTH2/CD294-PerCP-Cy5.5, 5 μl CD45-AmCyan, 5 μl CD34-PE, 5 μl CD11b-Pacific Blue (PB), 10 μl CD16-FITC (BD Biosciences), and 5 μl CD115-APC (R & D Systems Europe, Abingdon, United Kingdom).

    Techniques:

    Analytical validity of the assay demonstrated by analysis of the response of sorted viable CD45 + [CD3/CD19/CD20/CRTH2] − CD16 − CD115 − CD11b + CD34 + fibrocytes to stimulation with endothelin-1 (ET-1). (A) Representative flow cytometry analysis of the expression of intracellular α-smooth muscle actin (α-SMA) in freshly sorted and uncultured cells and in cells incubated for 6 days in culture medium alone or in culture medium supplemented with1 ng/ml ET-1. The black lines indicate staining with a specific anti α-SMA monoclonal antibody and the gray lines show nonspecific staining with the isotype control. (B) Quantitative analysis of the expression of intracellular α-SMA. The geometric mean fluorescence intensity (MFI) for α-SMA was divided by the corresponding value obtained with the isotype control. The graphs show individual geometric MFI ratio values obtained with the cells from 3 patients with controlled asthma (squares) and 3 patients with treatment-resistant asthma (circles) under each experimental condition. The horizontal lines indicate the means. AF, Alexa Fluor.

    Journal: BBA Clinical

    Article Title: Enumeration of circulating fibrocytes for clinical use in asthma by an optimized single-platform flow cytometry assay

    doi: 10.1016/j.bbacli.2014.06.002

    Figure Lengend Snippet: Analytical validity of the assay demonstrated by analysis of the response of sorted viable CD45 + [CD3/CD19/CD20/CRTH2] − CD16 − CD115 − CD11b + CD34 + fibrocytes to stimulation with endothelin-1 (ET-1). (A) Representative flow cytometry analysis of the expression of intracellular α-smooth muscle actin (α-SMA) in freshly sorted and uncultured cells and in cells incubated for 6 days in culture medium alone or in culture medium supplemented with1 ng/ml ET-1. The black lines indicate staining with a specific anti α-SMA monoclonal antibody and the gray lines show nonspecific staining with the isotype control. (B) Quantitative analysis of the expression of intracellular α-SMA. The geometric mean fluorescence intensity (MFI) for α-SMA was divided by the corresponding value obtained with the isotype control. The graphs show individual geometric MFI ratio values obtained with the cells from 3 patients with controlled asthma (squares) and 3 patients with treatment-resistant asthma (circles) under each experimental condition. The horizontal lines indicate the means. AF, Alexa Fluor.

    Article Snippet: One hundred μl of treated or untreated blood were added to a 5-ml round-bottom tube containing 2 μl of the viability dye 7-amino-actinomycin-D (7-AAD, BD Biosciences, San Jose, CA, USA), which is detected on the PerCP-Cy5.5 channel, and appropriate dilutions in stain buffer (BD Biosciences) of the following fluorochrome-labeled monoclonal antibodies (all mouse IgG1κ ): 2.5 μl CD3-PerCP-Cy5.5, 2.5 μl CD19-PerCP-Cy5.5, 2.5 μl CD20-PerCP-Cy5.5, 2.5 μl CRTH2/CD294-PerCP-Cy5.5, 5 μl CD45-AmCyan, 5 μl CD34-PE, 5 μl CD11b-Pacific Blue (PB), 10 μl CD16-FITC (BD Biosciences), and 5 μl CD115-APC (R & D Systems Europe, Abingdon, United Kingdom).

    Techniques: Flow Cytometry, Cytometry, Expressing, Incubation, Staining, Fluorescence

    Analytical validity of the assay demonstrated by analysis of the expression of type I collagen protein and gene in sorted viable CD45 + [CD3/CD19/CD20/CRTH2] − CD16 − CD115 − CD11b + CD34 + fibrocytes. (A) Representative flow cytometry analysis of the expression of intracellular type I collagen (COL1). The black line indicates staining with a specific anti-COL1 monoclonal antibody and the gray line shows nonspecific staining with the isotype control (B) Quantitative analysis of the expression of intracellular COL1. The geometric mean fluorescence intensity (MFI) for COL1 was divided by the corresponding value obtained with the isotype control. The graph shows individual geometric MFI ratio values obtained with the cells from 3 patients with controlled asthma (squares) and 3 patients with treatment-resistant asthma (circles). The horizontal lines indicate the means. (C) Analysis of the expression of the gene encoding the pro-α1 chain of COL1 (COL1A1) by reverse-transcription and real time polymerase chain reaction. The graph shows a representative example of quantification by using the relative standard curve method, where the mean threshold cycle (C T ) values from duplicate reactions are plotted against the log of the number of cells used in the reaction. The endogenous control was ACTB, the gene encoding β-actin. The normalized levels of expression of COL1A1 in sorted fibrocytes from 3 patients with treatment-resistant asthma were 0.255, 0.334 and 0.469 (∆C T method with efficiency correction).

    Journal: BBA Clinical

    Article Title: Enumeration of circulating fibrocytes for clinical use in asthma by an optimized single-platform flow cytometry assay

    doi: 10.1016/j.bbacli.2014.06.002

    Figure Lengend Snippet: Analytical validity of the assay demonstrated by analysis of the expression of type I collagen protein and gene in sorted viable CD45 + [CD3/CD19/CD20/CRTH2] − CD16 − CD115 − CD11b + CD34 + fibrocytes. (A) Representative flow cytometry analysis of the expression of intracellular type I collagen (COL1). The black line indicates staining with a specific anti-COL1 monoclonal antibody and the gray line shows nonspecific staining with the isotype control (B) Quantitative analysis of the expression of intracellular COL1. The geometric mean fluorescence intensity (MFI) for COL1 was divided by the corresponding value obtained with the isotype control. The graph shows individual geometric MFI ratio values obtained with the cells from 3 patients with controlled asthma (squares) and 3 patients with treatment-resistant asthma (circles). The horizontal lines indicate the means. (C) Analysis of the expression of the gene encoding the pro-α1 chain of COL1 (COL1A1) by reverse-transcription and real time polymerase chain reaction. The graph shows a representative example of quantification by using the relative standard curve method, where the mean threshold cycle (C T ) values from duplicate reactions are plotted against the log of the number of cells used in the reaction. The endogenous control was ACTB, the gene encoding β-actin. The normalized levels of expression of COL1A1 in sorted fibrocytes from 3 patients with treatment-resistant asthma were 0.255, 0.334 and 0.469 (∆C T method with efficiency correction).

    Article Snippet: One hundred μl of treated or untreated blood were added to a 5-ml round-bottom tube containing 2 μl of the viability dye 7-amino-actinomycin-D (7-AAD, BD Biosciences, San Jose, CA, USA), which is detected on the PerCP-Cy5.5 channel, and appropriate dilutions in stain buffer (BD Biosciences) of the following fluorochrome-labeled monoclonal antibodies (all mouse IgG1κ ): 2.5 μl CD3-PerCP-Cy5.5, 2.5 μl CD19-PerCP-Cy5.5, 2.5 μl CD20-PerCP-Cy5.5, 2.5 μl CRTH2/CD294-PerCP-Cy5.5, 5 μl CD45-AmCyan, 5 μl CD34-PE, 5 μl CD11b-Pacific Blue (PB), 10 μl CD16-FITC (BD Biosciences), and 5 μl CD115-APC (R & D Systems Europe, Abingdon, United Kingdom).

    Techniques: Expressing, Flow Cytometry, Cytometry, Staining, Fluorescence, Real-time Polymerase Chain Reaction

    Characterization of hMSCs. (A–C) Images of differentiated hMSCs after induction into specific tissues. (A) The lipid droplet accumulation in differentiated cells was visualized using Oil Red O staining after 2 weeks of adipogenic induction. (B) Calcium deposition was stained with Alizarin Red S after 2 weeks of osteogenic induction. (C) Glycosaminoglycans in cell pellets were revealed by Toluidine blue staining after 2 weeks of chondrogenic induction. (D) hMSCs (1×10 6 cells/ml) were stained with FITC- or PE- conjugated antibodies specific for human CD29, CD34, CD45, CD73, CD105, and HLA-DR.

    Journal: PLoS ONE

    Article Title: A p38 MAPK-Mediated Alteration of COX-2/PGE2 Regulates Immunomodulatory Properties in Human Mesenchymal Stem Cell Aging

    doi: 10.1371/journal.pone.0102426

    Figure Lengend Snippet: Characterization of hMSCs. (A–C) Images of differentiated hMSCs after induction into specific tissues. (A) The lipid droplet accumulation in differentiated cells was visualized using Oil Red O staining after 2 weeks of adipogenic induction. (B) Calcium deposition was stained with Alizarin Red S after 2 weeks of osteogenic induction. (C) Glycosaminoglycans in cell pellets were revealed by Toluidine blue staining after 2 weeks of chondrogenic induction. (D) hMSCs (1×10 6 cells/ml) were stained with FITC- or PE- conjugated antibodies specific for human CD29, CD34, CD45, CD73, CD105, and HLA-DR.

    Article Snippet: Flow cytometric analysis hMSCs were triturated into single cells and labeled with monoclonal mouse anti-human fluorochrome-conjugated antibodies: CD29-PE, CD34-FITC, CD45-FITC, CD105-FITC, CD73-PE, and HLA-DR (BD Bioscience, San Jose, CA).

    Techniques: Staining

    Multicolour flow cytometric immunophenotypic analysis with monoclonal antibodies. Intraoral MSPCs at passages 3–7 were labelled with specific fluorochrome-conjugated monoclonal antibodies against the indicated surface antigens and analysed by flow cytometry. Analysed MSPCs demonstrated the same immunophenotype, with expression of (a) CD73, (b) CD90, and (c) CD105 but no expression of (d) CD14, (e) CD19, (f) CD34, (g) CD45, and (h) HLA-DR. A representative example of ten experiments is shown. PE: phycoerythrin; APC: allophycocyanin; PerCP: peridium-chlorophyll protein complex; FITC: fluorescein isothiocyanate.

    Journal: BioMed Research International

    Article Title: Effect of Cyclic Mechanical Stimulation on the Expression of Osteogenesis Genes in Human Intraoral Mesenchymal Stromal and Progenitor Cells

    doi: 10.1155/2014/189516

    Figure Lengend Snippet: Multicolour flow cytometric immunophenotypic analysis with monoclonal antibodies. Intraoral MSPCs at passages 3–7 were labelled with specific fluorochrome-conjugated monoclonal antibodies against the indicated surface antigens and analysed by flow cytometry. Analysed MSPCs demonstrated the same immunophenotype, with expression of (a) CD73, (b) CD90, and (c) CD105 but no expression of (d) CD14, (e) CD19, (f) CD34, (g) CD45, and (h) HLA-DR. A representative example of ten experiments is shown. PE: phycoerythrin; APC: allophycocyanin; PerCP: peridium-chlorophyll protein complex; FITC: fluorescein isothiocyanate.

    Article Snippet: The commercial monoclonal antibodies CD73 PE, CD90 APC, CD105 PE, CD45 APC-Cy7, CD34 APC, CD14 FITC, CD19 APC, and HLA-DR APC (BD Bioscience, San Jose, CA) were applied for characterization.

    Techniques: Flow Cytometry, Cytometry, Expressing

    The tumor cells tested positive for alpha smooth muscle actin, calponin, H-caldesmon, but were negative for CD34, p53, estrogen receptor, progesterone receptor, and Epstein-Barr virus. In approximately 25% of all the tumor cells, the Ki67 proliferative index was positive. A: Pathological finding of thyroid gland stained (10 ×); B: Positive immunohistochemical staining for α-smooth muscle actin (10 ×); C: Negative immunohistochemical staining for P53 (20 ×); D: Pathological finding of 25% positive Ki67 index in tumor cells (20 ×); E: Negative immunohistochemical staining for CD34 (10 ×); F: Positive immunohistochemical staining for calponin (20 ×).

    Journal: World Journal of Clinical Cases

    Article Title: Primary leiomyosarcoma of the thyroid gland with prior malignancy and radiotherapy: A case report and review of literature

    doi: 10.12998/wjcc.v7.i4.473

    Figure Lengend Snippet: The tumor cells tested positive for alpha smooth muscle actin, calponin, H-caldesmon, but were negative for CD34, p53, estrogen receptor, progesterone receptor, and Epstein-Barr virus. In approximately 25% of all the tumor cells, the Ki67 proliferative index was positive. A: Pathological finding of thyroid gland stained (10 ×); B: Positive immunohistochemical staining for α-smooth muscle actin (10 ×); C: Negative immunohistochemical staining for P53 (20 ×); D: Pathological finding of 25% positive Ki67 index in tumor cells (20 ×); E: Negative immunohistochemical staining for CD34 (10 ×); F: Positive immunohistochemical staining for calponin (20 ×).

    Article Snippet: Immunohistochemically, the tumor cells tested positive for alpha SMA, calponin, and H-caldesmon, but were negative for CD34, p53, estrogen receptor, progesterone receptor, and EBV.

    Techniques: Staining, Immunohistochemistry

    Lineage Tracing of Inter-Follicular Epidermis Stem Cells in Aspm-CreER Mice (A) Scheme of the epithelial skin structure with epidermis at the top and HF at the bottom. CL, cornified layer; G/SL, granular/spinous layer; BL, basal layer; SG, sebaceous gland; HFSC, HF stem cell; HG, hair germ; DP, dermal papilla. Note the marking of epidermis versus HF using Aspm -CreER versus Lgr5 -CreER genetic drivers. (B) Gene expression analysis of FACS-sorted inter-follicular epidermis (IFE) BL cells and HFSCs in Catagen (PD41) and Anagen (PD24). N = 3 of biological replicates; ∗ p = 0.009, ∗∗ p = 0.0002, ∗∗∗ p = 0.017, ∗∗∗∗ p = 0.001. HFSCs are FACS sorted as α6-integrin+/CD34+ and BL cells were α6-integrin+/CD34− cells. Student’s t test was used for all significance tests. (C) Scheme of lineage tracing using Aspm -CreER;Rosa26-tdTomato mice after tamoxifen (TM) injection (red arrows) and different experimental endpoints (black arrowhead); 'sac' stands for time of sacrifice. (D) Back skin of Aspm -CreER;tdTomato mice treated as shown in (C) at endpoints indicated. Arrowhead shows Aspm -CreER-marked cells positive for tdTomato. Asterisk indicates autofluorescence from the hair shaft. N = 2 for TM−; N = 3 for the rest of the time points. (E) Experimental scheme and whole-mouse pictures showing back skin hair plucking of Aspm -CreER;tdTomato mice at time points indicated. Note that the right-hand side was plucked, and the hairs grew back by 2 weeks post-plucking. N = 3. (F) Lineage tracing of Aspm -CreER-marked positive IFE cells at PD98, 1 month after plucking. Left panel shows no migration in the HFs. Middle panel shows migration of Aspm cells to the infundibulum (arrowhead). Right panel shows migration of Aspm cells to the bulge (arrowhead). N = 3. The table was generated from averaging 100 HFs from 3 mice at 1 month after hair plucking. Scale bars, 20 μm.

    Journal: Stem Cell Reports

    Article Title: Histone H3 K4/9/27 Trimethylation Levels Affect Wound Healing and Stem Cell Dynamics in Adult Skin

    doi: 10.1016/j.stemcr.2019.11.007

    Figure Lengend Snippet: Lineage Tracing of Inter-Follicular Epidermis Stem Cells in Aspm-CreER Mice (A) Scheme of the epithelial skin structure with epidermis at the top and HF at the bottom. CL, cornified layer; G/SL, granular/spinous layer; BL, basal layer; SG, sebaceous gland; HFSC, HF stem cell; HG, hair germ; DP, dermal papilla. Note the marking of epidermis versus HF using Aspm -CreER versus Lgr5 -CreER genetic drivers. (B) Gene expression analysis of FACS-sorted inter-follicular epidermis (IFE) BL cells and HFSCs in Catagen (PD41) and Anagen (PD24). N = 3 of biological replicates; ∗ p = 0.009, ∗∗ p = 0.0002, ∗∗∗ p = 0.017, ∗∗∗∗ p = 0.001. HFSCs are FACS sorted as α6-integrin+/CD34+ and BL cells were α6-integrin+/CD34− cells. Student’s t test was used for all significance tests. (C) Scheme of lineage tracing using Aspm -CreER;Rosa26-tdTomato mice after tamoxifen (TM) injection (red arrows) and different experimental endpoints (black arrowhead); 'sac' stands for time of sacrifice. (D) Back skin of Aspm -CreER;tdTomato mice treated as shown in (C) at endpoints indicated. Arrowhead shows Aspm -CreER-marked cells positive for tdTomato. Asterisk indicates autofluorescence from the hair shaft. N = 2 for TM−; N = 3 for the rest of the time points. (E) Experimental scheme and whole-mouse pictures showing back skin hair plucking of Aspm -CreER;tdTomato mice at time points indicated. Note that the right-hand side was plucked, and the hairs grew back by 2 weeks post-plucking. N = 3. (F) Lineage tracing of Aspm -CreER-marked positive IFE cells at PD98, 1 month after plucking. Left panel shows no migration in the HFs. Middle panel shows migration of Aspm cells to the infundibulum (arrowhead). Right panel shows migration of Aspm cells to the bulge (arrowhead). N = 3. The table was generated from averaging 100 HFs from 3 mice at 1 month after hair plucking. Scale bars, 20 μm.

    Article Snippet: The cell suspensions were labeled with anti-CD34-Biotin (1:50, no. 13-0341-85, eBioscience), α-Streptavidin-APC (1:100, no. 554067, BD Biosciences) and anti-α6-integrin-PE (1:40, CD49f, no. 555736, BD Biosciences) to isolate HFSCs (CD34+, α6-integrin+) and BL cells (CD34−, α6-integrin+).

    Techniques: Mouse Assay, Expressing, FACS, Injection, Migration, Generated

    BMP4 Expression Is Increased by Blocking Hypomethylation at Late Anagen/Catagen (A) Intensity graph of BMP4 normalized by β-actin of CT-treated (blue) or DI-treated (red) mice at late Anagen/Catagen extracted from western blots in Figure S5 A at the ages and stages indicated. N = 2 for PD35; N = 3 for other ages. A, Anagen; C, Catagen; T, Telogen. Corresponding histone methylation levels normalized by H3 of CT-treated mice total skin at various ages are shown in gray and were extracted from Lee et al. (2016) . Note the decrease in histone methylation level from Anagen to Catagen transition, accompanied by an increase in BMP4 levels. From Catagen to Telogen, histone methylation and BMP4 levels positively correlate, while from Telogen to Anagen, methylation continues to increase while BMP4 levels decrease. (B) Gene expression analysis of Bmp4 mRNA in CT- or DI-treated total back skin. N = 3 of biological replicates; ∗ p = 0.018. (C) Skin sections immunostained with BMP4 show expression of BMP4 in bulge and epidermis with apparent increase in DI-treated skin. Bu, bulge; Ep, epidermis. (D) BMP4 fluorescence signal quantification in the bulge (CD34+ cells) and the basal layer cells as shown in Figure S5B. N = 3; ∗ p = 0.022, ∗∗ p = 0.003. Bu, bulge; Ep, epidermis. (E) Scheme of the Bmp4 promoter region with three primer sets upstream of the transcription start site (TSS) used for the P experiment of CT- or DI-treated skin. (F) ChIP signal fold change of Bmp4 promoter regions 1, 2, and 3 shown in (E). ∗ p = 0.047, ∗∗ p = 0.012, ∗∗∗ p = 0.029, ∗∗∗∗ p = 0.017, ∗∗∗∗∗ p = 0.04; N = 2 for Anagen (PD35) samples, N = 3 for Catagen (PD39) samples. The samples that were used for the ChIP experiments were the total skin samples of biological replicates after applying DI or control vehicles (CT). (G) Scheme of the NOGGIN-coated bead injection experiment after applying DI. (H) Skin sections immunostained with CD34 and Ki67 from mice sacrificed at PD53 after injecting NOGGIN in DI-treated skin. (I) Quantification of the HFs in each hair cycle from (H). N = 3; ∗ p = 0.014. (J) Skin sections immunostained with CD34 and pSMAD1, 5, 9 at PD49 after applying DI at PD35-42. (K) Quantification of fluorescence signals of pSMAD1, 5, 9 in the bulge (CD34+ cells) as shown in (J). N = 3; ∗ p = 0.004. (L) Working model describing histone methylation level crosstalk with BMP4. Student’s t test was used for all significance test. All experiments were executed twice. Scale bars, 20 μm.

    Journal: Stem Cell Reports

    Article Title: Histone H3 K4/9/27 Trimethylation Levels Affect Wound Healing and Stem Cell Dynamics in Adult Skin

    doi: 10.1016/j.stemcr.2019.11.007

    Figure Lengend Snippet: BMP4 Expression Is Increased by Blocking Hypomethylation at Late Anagen/Catagen (A) Intensity graph of BMP4 normalized by β-actin of CT-treated (blue) or DI-treated (red) mice at late Anagen/Catagen extracted from western blots in Figure S5 A at the ages and stages indicated. N = 2 for PD35; N = 3 for other ages. A, Anagen; C, Catagen; T, Telogen. Corresponding histone methylation levels normalized by H3 of CT-treated mice total skin at various ages are shown in gray and were extracted from Lee et al. (2016) . Note the decrease in histone methylation level from Anagen to Catagen transition, accompanied by an increase in BMP4 levels. From Catagen to Telogen, histone methylation and BMP4 levels positively correlate, while from Telogen to Anagen, methylation continues to increase while BMP4 levels decrease. (B) Gene expression analysis of Bmp4 mRNA in CT- or DI-treated total back skin. N = 3 of biological replicates; ∗ p = 0.018. (C) Skin sections immunostained with BMP4 show expression of BMP4 in bulge and epidermis with apparent increase in DI-treated skin. Bu, bulge; Ep, epidermis. (D) BMP4 fluorescence signal quantification in the bulge (CD34+ cells) and the basal layer cells as shown in Figure S5B. N = 3; ∗ p = 0.022, ∗∗ p = 0.003. Bu, bulge; Ep, epidermis. (E) Scheme of the Bmp4 promoter region with three primer sets upstream of the transcription start site (TSS) used for the P experiment of CT- or DI-treated skin. (F) ChIP signal fold change of Bmp4 promoter regions 1, 2, and 3 shown in (E). ∗ p = 0.047, ∗∗ p = 0.012, ∗∗∗ p = 0.029, ∗∗∗∗ p = 0.017, ∗∗∗∗∗ p = 0.04; N = 2 for Anagen (PD35) samples, N = 3 for Catagen (PD39) samples. The samples that were used for the ChIP experiments were the total skin samples of biological replicates after applying DI or control vehicles (CT). (G) Scheme of the NOGGIN-coated bead injection experiment after applying DI. (H) Skin sections immunostained with CD34 and Ki67 from mice sacrificed at PD53 after injecting NOGGIN in DI-treated skin. (I) Quantification of the HFs in each hair cycle from (H). N = 3; ∗ p = 0.014. (J) Skin sections immunostained with CD34 and pSMAD1, 5, 9 at PD49 after applying DI at PD35-42. (K) Quantification of fluorescence signals of pSMAD1, 5, 9 in the bulge (CD34+ cells) as shown in (J). N = 3; ∗ p = 0.004. (L) Working model describing histone methylation level crosstalk with BMP4. Student’s t test was used for all significance test. All experiments were executed twice. Scale bars, 20 μm.

    Article Snippet: The cell suspensions were labeled with anti-CD34-Biotin (1:50, no. 13-0341-85, eBioscience), α-Streptavidin-APC (1:100, no. 554067, BD Biosciences) and anti-α6-integrin-PE (1:40, CD49f, no. 555736, BD Biosciences) to isolate HFSCs (CD34+, α6-integrin+) and BL cells (CD34−, α6-integrin+).

    Techniques: Expressing, Blocking Assay, Mouse Assay, Western Blot, Methylation, Fluorescence, Chromatin Immunoprecipitation, Injection

    In vivo inhibition of miR-138-5p reduces the pulmonary vascular cell proliferation in MCT-PH rats. a Immunofluorescence staining of frozen rat lung sections and confocal imaging with Click-iT 5-ethynyl-2′-deoxyuridine (EdU; white nuclei = EdU-positive nuclei = proliferating cells, yellow arrow) in combination with α-smooth muscle actin (α-SMA; in green, CD34; in red). Scale bar = 20 μm. The cells were counterstained with DAPI (blue). b Quantification of the percentage of proliferating cells in the lungs. ( n = 6 different rats per condition). * P

    Journal: Respiratory Research

    Article Title: In vivo miR-138-5p inhibition alleviates monocrotaline-induced pulmonary hypertension and normalizes pulmonary KCNK3 and SLC45A3 expression

    doi: 10.1186/s12931-020-01444-7

    Figure Lengend Snippet: In vivo inhibition of miR-138-5p reduces the pulmonary vascular cell proliferation in MCT-PH rats. a Immunofluorescence staining of frozen rat lung sections and confocal imaging with Click-iT 5-ethynyl-2′-deoxyuridine (EdU; white nuclei = EdU-positive nuclei = proliferating cells, yellow arrow) in combination with α-smooth muscle actin (α-SMA; in green, CD34; in red). Scale bar = 20 μm. The cells were counterstained with DAPI (blue). b Quantification of the percentage of proliferating cells in the lungs. ( n = 6 different rats per condition). * P

    Article Snippet: To characterize the EdU-positive cells, we performed double-immunostaining with FITC-labelled anti-α-SMA (clone 1A4, Sigma-Aldrich, Lyon, France, 1/100) or with anti-CD34 (EP373Y, Abcam, Amsterdam, 1/200).

    Techniques: In Vivo, Inhibition, Immunofluorescence, Staining, Imaging

    After gating for exclusion of debris and isolation of single cells, CD45-/CD34+/CD31- cells were isolated (21.9 ±± 2.4%). Flow cytometry gates were drawn based on unstained control ASCs.

    Journal: Plastic and reconstructive surgery

    Article Title: Purified Adipose Derived Stromal Cells Provide Superior Fat Graft Retention Compared to Unenriched Stromal Vascular Fraction

    doi: 10.1097/PRS.0000000000003165

    Figure Lengend Snippet: After gating for exclusion of debris and isolation of single cells, CD45-/CD34+/CD31- cells were isolated (21.9 ±± 2.4%). Flow cytometry gates were drawn based on unstained control ASCs.

    Article Snippet: The remaining cells were washed in staining buffer (1% fetal bovine serum, 1% P188, 1% penicillin-streptomycin) prior to resuspension in a solution containing anti-CD45, anti-CD34, and anti-CD31 antibodies (eBioscience, Inc., San Diego, CA).

    Techniques: Isolation, Flow Cytometry, Cytometry

    Morphology and phenotype of CD40-activated CD34 + HPC and in situ localization of CD1a − /CD40 − , DR + DC. CD34 + HPC prepared as described were seeded onto control (CD32L cells) cultures ( a ) or onto CD40L (CD40L-L cells) cultures ( b–i ) and evaluated at days 8 ( a and b ) and days 14 ( c – i ). Photomicrographs were taken from cells in culture wells ( a–d ) at ×200 ( a and b ) and ×400 ( c and d ) magnification, and from cytospin preparations ( e ) or from BioRad slides ( f–i ) at ×1,000 magnification. In ( c ) and ( d ), the same field was taken at a 15-s interval to show dendrite motility. ( e ) May– Grünwald staining of DC. ( f ) HIV–gp120 binding. ( g ) shows a CD26 + DC. ( h ) A rel-B + DC together with a rel-B − , nondendritic cell. ( i ) MHC class II DR + DC. ( j and k ) Tonsil sections stained for CD1a and CD40 (both in blue ) and MHC class II DR ( red ) showing single DR + , CD1a − and CD40 − DC scattered in the T cell area. ( j ) ×400 magnification and ( k ) ×1,000 magnification.

    Journal: The Journal of Experimental Medicine

    Article Title: CD40 Ligation on Human Cord Blood CD34+Hematopoietic Progenitors Induces Their Proliferation and Differentiation into Functional Dendritic Cells

    doi:

    Figure Lengend Snippet: Morphology and phenotype of CD40-activated CD34 + HPC and in situ localization of CD1a − /CD40 − , DR + DC. CD34 + HPC prepared as described were seeded onto control (CD32L cells) cultures ( a ) or onto CD40L (CD40L-L cells) cultures ( b–i ) and evaluated at days 8 ( a and b ) and days 14 ( c – i ). Photomicrographs were taken from cells in culture wells ( a–d ) at ×200 ( a and b ) and ×400 ( c and d ) magnification, and from cytospin preparations ( e ) or from BioRad slides ( f–i ) at ×1,000 magnification. In ( c ) and ( d ), the same field was taken at a 15-s interval to show dendrite motility. ( e ) May– Grünwald staining of DC. ( f ) HIV–gp120 binding. ( g ) shows a CD26 + DC. ( h ) A rel-B + DC together with a rel-B − , nondendritic cell. ( i ) MHC class II DR + DC. ( j and k ) Tonsil sections stained for CD1a and CD40 (both in blue ) and MHC class II DR ( red ) showing single DR + , CD1a − and CD40 − DC scattered in the T cell area. ( j ) ×400 magnification and ( k ) ×1,000 magnification.

    Article Snippet: However, at 24 h (data not shown), the percentage of cycling cells in CD40L cultures was only marginally higher than that of control cultures, indicating that CD40L probably recruits a small proportion of CD34+ cells whose cycling progeny accumulates with time.

    Techniques: In Situ, Staining, Binding Assay

    CD40 ligation induces proliferation of human CD34 + progenitor cells. ( A ) Kinetics, ( B , E ) specificity, ( C , D ) cell cycle analysis. CD34HPC cultured as described in Materials and Methods were either pulsed overnight with 1 μCi of [ 3 H]thymidine ( A , B ) or counted by Trypan blue exclusion ( E ). Experiments shown are representative of three experiments and data are presented as mean cpm ± SD ( A and B ) and cell numbers per ml ± SD ( E ), as determined in triplicate wells. For quantitating recovery of viable cells, 10 5 progenitor cells (indicated by horizontal line in medium control bar) were seeded in 1.0 ml of culture medium onto 10 5 CD40L + cells or control CD32 cells. Results were evaluated at days 4, 8, and 15 ( A ) or at day 8 only ( B , E ). Where indicated, cultures were supplemented with mAbs to CD40 (mAb 89), CD40L (mAb LL48), or isotype-matched control antibodies. ( C , D ) Cell cycle analysis. CD34HPC freshly isolated (data not shown) or cultured either on CD32L ( C ) or CD40L-transfected fibroblasts ( D ) for 3 d were resuspended and incubated with Hoechst 33342. To quantitate specifically CD34HPC entering into cycle, Hoechst-stained cells were subsequently labeled with a mAb to CD34, washed, and analyzed with a FACSTAR +® flow cytometer.

    Journal: The Journal of Experimental Medicine

    Article Title: CD40 Ligation on Human Cord Blood CD34+Hematopoietic Progenitors Induces Their Proliferation and Differentiation into Functional Dendritic Cells

    doi:

    Figure Lengend Snippet: CD40 ligation induces proliferation of human CD34 + progenitor cells. ( A ) Kinetics, ( B , E ) specificity, ( C , D ) cell cycle analysis. CD34HPC cultured as described in Materials and Methods were either pulsed overnight with 1 μCi of [ 3 H]thymidine ( A , B ) or counted by Trypan blue exclusion ( E ). Experiments shown are representative of three experiments and data are presented as mean cpm ± SD ( A and B ) and cell numbers per ml ± SD ( E ), as determined in triplicate wells. For quantitating recovery of viable cells, 10 5 progenitor cells (indicated by horizontal line in medium control bar) were seeded in 1.0 ml of culture medium onto 10 5 CD40L + cells or control CD32 cells. Results were evaluated at days 4, 8, and 15 ( A ) or at day 8 only ( B , E ). Where indicated, cultures were supplemented with mAbs to CD40 (mAb 89), CD40L (mAb LL48), or isotype-matched control antibodies. ( C , D ) Cell cycle analysis. CD34HPC freshly isolated (data not shown) or cultured either on CD32L ( C ) or CD40L-transfected fibroblasts ( D ) for 3 d were resuspended and incubated with Hoechst 33342. To quantitate specifically CD34HPC entering into cycle, Hoechst-stained cells were subsequently labeled with a mAb to CD34, washed, and analyzed with a FACSTAR +® flow cytometer.

    Article Snippet: However, at 24 h (data not shown), the percentage of cycling cells in CD40L cultures was only marginally higher than that of control cultures, indicating that CD40L probably recruits a small proportion of CD34+ cells whose cycling progeny accumulates with time.

    Techniques: Ligation, Cell Cycle Assay, Cell Culture, Isolation, Transfection, Incubation, Staining, Labeling, Flow Cytometry, Cytometry

    CD40 triggering of CD34 + progenitors specifically induces the generation of DC that accumulate with time. ( A ) Human CD34 + progenitor cells seeded in CD40L (LCD40L cells) or in control cultures (LCD32 cells) were evaluated for DC generation at different times. ( B ) The effect of neutralizing antibodies to CD40, CD40L, and GM–CSF, or an irrelevant control Ab was also determined at day 14. Results are expressed as percentage DC ± SD evaluated in triplicates. Cultures and evaluation of DC production were performed as described in Materials and Methods.

    Journal: The Journal of Experimental Medicine

    Article Title: CD40 Ligation on Human Cord Blood CD34+Hematopoietic Progenitors Induces Their Proliferation and Differentiation into Functional Dendritic Cells

    doi:

    Figure Lengend Snippet: CD40 triggering of CD34 + progenitors specifically induces the generation of DC that accumulate with time. ( A ) Human CD34 + progenitor cells seeded in CD40L (LCD40L cells) or in control cultures (LCD32 cells) were evaluated for DC generation at different times. ( B ) The effect of neutralizing antibodies to CD40, CD40L, and GM–CSF, or an irrelevant control Ab was also determined at day 14. Results are expressed as percentage DC ± SD evaluated in triplicates. Cultures and evaluation of DC production were performed as described in Materials and Methods.

    Article Snippet: However, at 24 h (data not shown), the percentage of cycling cells in CD40L cultures was only marginally higher than that of control cultures, indicating that CD40L probably recruits a small proportion of CD34+ cells whose cycling progeny accumulates with time.

    Techniques:

    Design and Validation of a Unified Vector Encoding iMC and PSCA.ζ CAR (A) Retroviral transgene design encoding MyD88, CD40, tandem FKBP12v36, T2A, signal peptide, A11 scFv variable light (V L ) and variable heavy (V H ) domains, the minimal CD34 epitope, CD8α stalk and transmembrane region, and the CD3ζ signaling domain. (B) Flow cytometric analysis to determine transduction efficiency using anti-CD3 and anti-CD34 antibodies compared to non-transduced (NT) T cells. (C) iMC-PSCA.ζ-modified T cells only proliferate in the presence of exogenous IL-2 (100 U/mL) when stimulated with 10 nM rimiducid (Rim), but they demonstrate antigen-independent survival in the presence of IL-2 or with weekly iMC activation over 5 weeks (two individual donors shown). (D) T cells were transduced with EGFPluc or with both EGFPluc and iMC-PSCA.ζ-encoding-retrovirus and subsequently cultured with HPAC-RFP tumor cells at a 1:10 effector-to-tumor (E:T) cell ratio for 48 hr with variable Rim concentrations (0–250 nM) (n = 3). IL-6 and IL-2 production was measured by ELISA. (E) Control T cells (EGFPluc) or iMC-PSCA.ζ/EGFPluc-modified T cells were cultured at a 1:10 E:T ratio with HPAC-RFP cells at variable Rim concentrations (0–250 nM) for 7 days. Tumor cell killing (RFP; red) and T cell (EGFP; green) proliferation were measured by live-cell imaging using an IncuCyte imager. (F and G) EGFPluc and iMC-PSCA.ζ/EGFPluc-modified T cells were cultured with HPAC-RFP tumor cells at an E:T ratio of 1:20 and imaged over 7 days (n = 2). T cells transduced with iMC-PSCA.ζ/EGFPluc were stimulated with or without 2 nM Rim on day 0 of culture initiation.

    Journal: Molecular Therapy

    Article Title: Regulated Expansion and Survival of Chimeric Antigen Receptor-Modified T Cells Using Small Molecule-Dependent Inducible MyD88/CD40

    doi: 10.1016/j.ymthe.2017.06.014

    Figure Lengend Snippet: Design and Validation of a Unified Vector Encoding iMC and PSCA.ζ CAR (A) Retroviral transgene design encoding MyD88, CD40, tandem FKBP12v36, T2A, signal peptide, A11 scFv variable light (V L ) and variable heavy (V H ) domains, the minimal CD34 epitope, CD8α stalk and transmembrane region, and the CD3ζ signaling domain. (B) Flow cytometric analysis to determine transduction efficiency using anti-CD3 and anti-CD34 antibodies compared to non-transduced (NT) T cells. (C) iMC-PSCA.ζ-modified T cells only proliferate in the presence of exogenous IL-2 (100 U/mL) when stimulated with 10 nM rimiducid (Rim), but they demonstrate antigen-independent survival in the presence of IL-2 or with weekly iMC activation over 5 weeks (two individual donors shown). (D) T cells were transduced with EGFPluc or with both EGFPluc and iMC-PSCA.ζ-encoding-retrovirus and subsequently cultured with HPAC-RFP tumor cells at a 1:10 effector-to-tumor (E:T) cell ratio for 48 hr with variable Rim concentrations (0–250 nM) (n = 3). IL-6 and IL-2 production was measured by ELISA. (E) Control T cells (EGFPluc) or iMC-PSCA.ζ/EGFPluc-modified T cells were cultured at a 1:10 E:T ratio with HPAC-RFP cells at variable Rim concentrations (0–250 nM) for 7 days. Tumor cell killing (RFP; red) and T cell (EGFP; green) proliferation were measured by live-cell imaging using an IncuCyte imager. (F and G) EGFPluc and iMC-PSCA.ζ/EGFPluc-modified T cells were cultured with HPAC-RFP tumor cells at an E:T ratio of 1:20 and imaged over 7 days (n = 2). T cells transduced with iMC-PSCA.ζ/EGFPluc were stimulated with or without 2 nM Rim on day 0 of culture initiation.

    Article Snippet: Phenotypic analysis of T cells isolated from animals was assessed by nine-color flow cytometry using the following antibody panel: anti-murine CD45-BV510, anti-CD3-APC/Cy7, anti-CD4-PerCP-Cy5.5, anti-CD8-BV711, anti-CD34-PE, anti-CD45RA-fluorescein isothiocyanate (FITC), anti-CD62L-APC, anti-CD95-ECD, and anti-PD-1-BV421 (BD Biosciences).

    Techniques: Plasmid Preparation, Flow Cytometry, Transduction, Modification, Activation Assay, Cell Culture, Enzyme-linked Immunosorbent Assay, Live Cell Imaging

    iMC Enhances Potency of GD2-Targeted CARs Compared to Conventional Costimulatory Molecules (A) Vector design for GD2 CAR-T cells using various costimulatory signaling domains. (B) Transduction efficiency of GD2 CAR-T cells (n = 4) as measured by CD3 + CD34 + expression by flow cytometry. (C and D) IncuCyte coculture of CAR-modified T cells with GD2 + ).

    Journal: Molecular Therapy

    Article Title: Regulated Expansion and Survival of Chimeric Antigen Receptor-Modified T Cells Using Small Molecule-Dependent Inducible MyD88/CD40

    doi: 10.1016/j.ymthe.2017.06.014

    Figure Lengend Snippet: iMC Enhances Potency of GD2-Targeted CARs Compared to Conventional Costimulatory Molecules (A) Vector design for GD2 CAR-T cells using various costimulatory signaling domains. (B) Transduction efficiency of GD2 CAR-T cells (n = 4) as measured by CD3 + CD34 + expression by flow cytometry. (C and D) IncuCyte coculture of CAR-modified T cells with GD2 + ).

    Article Snippet: Phenotypic analysis of T cells isolated from animals was assessed by nine-color flow cytometry using the following antibody panel: anti-murine CD45-BV510, anti-CD3-APC/Cy7, anti-CD4-PerCP-Cy5.5, anti-CD8-BV711, anti-CD34-PE, anti-CD45RA-fluorescein isothiocyanate (FITC), anti-CD62L-APC, anti-CD95-ECD, and anti-PD-1-BV421 (BD Biosciences).

    Techniques: Plasmid Preparation, Transduction, Expressing, Flow Cytometry, Cytometry, Modification

    iMC Enhances Proliferation and Anti-tumor Efficacy of CD123-Targeted CARs against Leukemia (A) Transduction efficiency as measured by CD3 + CD34 + expression in non-transduced and iMC-CD123.ζ-modified T cells. (B) Flow cytometric analysis of CD123 expression on four leukemia and lymphoma cell lines (KG-1, MOLM-13, THP-1, and U-937). (C) Non-transduced (NT) and iMC-CD123.ζ-modified T cells were cultured together with CD123 + (KG-1, MOLM-13, or THP-1) or CD123 − (U-937) cancer cell lines for 7 days at an E:T ratio of 1:1 effector-to-target cells and evaluated for outgrowth of GFP + tumor cells. (D) T cell proliferation of NT and iMC-CD123.ζ-modified T cells following coculture with KG-1, MOLM-13, THP-1, and U-937 cell lines was calculated as the fold increase following costimulation with 1 nM Rim. (E) IL-2 production was measured by ELISA following 48 hr of coculture of NT or iMC-CD123.ζ-modified T cells with THP-1 cells in the presence or absence of 1 nM Rim. (F) NSG mice (n = 5 per group) were challenged with 1 × 10 6 THP-1-EGFPluc tumor cells via i.v. injection, followed by i.v. treatment with 2.5 × 10 6 ). *p

    Journal: Molecular Therapy

    Article Title: Regulated Expansion and Survival of Chimeric Antigen Receptor-Modified T Cells Using Small Molecule-Dependent Inducible MyD88/CD40

    doi: 10.1016/j.ymthe.2017.06.014

    Figure Lengend Snippet: iMC Enhances Proliferation and Anti-tumor Efficacy of CD123-Targeted CARs against Leukemia (A) Transduction efficiency as measured by CD3 + CD34 + expression in non-transduced and iMC-CD123.ζ-modified T cells. (B) Flow cytometric analysis of CD123 expression on four leukemia and lymphoma cell lines (KG-1, MOLM-13, THP-1, and U-937). (C) Non-transduced (NT) and iMC-CD123.ζ-modified T cells were cultured together with CD123 + (KG-1, MOLM-13, or THP-1) or CD123 − (U-937) cancer cell lines for 7 days at an E:T ratio of 1:1 effector-to-target cells and evaluated for outgrowth of GFP + tumor cells. (D) T cell proliferation of NT and iMC-CD123.ζ-modified T cells following coculture with KG-1, MOLM-13, THP-1, and U-937 cell lines was calculated as the fold increase following costimulation with 1 nM Rim. (E) IL-2 production was measured by ELISA following 48 hr of coculture of NT or iMC-CD123.ζ-modified T cells with THP-1 cells in the presence or absence of 1 nM Rim. (F) NSG mice (n = 5 per group) were challenged with 1 × 10 6 THP-1-EGFPluc tumor cells via i.v. injection, followed by i.v. treatment with 2.5 × 10 6 ). *p

    Article Snippet: Phenotypic analysis of T cells isolated from animals was assessed by nine-color flow cytometry using the following antibody panel: anti-murine CD45-BV510, anti-CD3-APC/Cy7, anti-CD4-PerCP-Cy5.5, anti-CD8-BV711, anti-CD34-PE, anti-CD45RA-fluorescein isothiocyanate (FITC), anti-CD62L-APC, anti-CD95-ECD, and anti-PD-1-BV421 (BD Biosciences).

    Techniques: Transduction, Expressing, Modification, Flow Cytometry, Cell Culture, Enzyme-linked Immunosorbent Assay, Mouse Assay, Injection

    Immunohistochemical features of PLNTY. Strong labeling for GFAP as seen in case 3 ( a ), OLIG2 positivity as seen in case 2 ( b ), negativity for HuC/HuD as seen in case 6, and expression of BRAF V600E as seen in case 3. Diffuse strong expression of CD34 in both tumor cells ( e , f ) and peripherally associated ramified neural elements ( g , h ) as seen in case 3 ( e , g ) and case 6 ( f , h )

    Journal: Acta Neuropathologica

    Article Title: Polymorphous low-grade neuroepithelial tumor of the young (PLNTY): an epileptogenic neoplasm with oligodendroglioma-like components, aberrant CD34 expression, and genetic alterations involving the MAP kinase pathway

    doi: 10.1007/s00401-016-1639-9

    Figure Lengend Snippet: Immunohistochemical features of PLNTY. Strong labeling for GFAP as seen in case 3 ( a ), OLIG2 positivity as seen in case 2 ( b ), negativity for HuC/HuD as seen in case 6, and expression of BRAF V600E as seen in case 3. Diffuse strong expression of CD34 in both tumor cells ( e , f ) and peripherally associated ramified neural elements ( g , h ) as seen in case 3 ( e , g ) and case 6 ( f , h )

    Article Snippet: While the immunohistochemical labeling of some glioblastomas for CD34 is recognized [ , ], this phenomenon is generally foreign to infiltrating astrocytomas of lower grade and to neoplasms of the oligodendroglioma group [ , , , , – , , , , , , – ].

    Techniques: Immunohistochemistry, Labeling, Expressing

    Fibrocyte recruitment to kidney and spleen in response to ureteral obstruction. Male coll-GFP bone marrow was transplanted into female WT lethally irradiated mice. Chimerism was confirmed at 6 weeks by assessing leukocytes for Y chromosome. A: Image of perivascular GFP+ cells in d7 UUO kidney from coll-GFP ⇒WT bone marrow chimeric mice (magnification = original ×400). B: Confirmation of extensive fibrosis in this model by picrosirius red staining of d14 UUO kidney from chimeric mice. Note prominent perivascular fibrosis ( arrows ) as well as interstitial fibrosis. C–D: Image of d7 in UUO kidney from coll-GFP ⇒WT BM chimeric mice showing GFP+ cells co-expressing CD34 and CD45, confirming these cells to be fibrocytes (magnification = original ×400). E–F: Image of d7 UUO kidney showing BM-derived coll1a1 -GFP+ cells lacking αSMA or S100A4 expression (magnification = original ×400). G: Red pulp of spleen from coll-GFP ⇒WT BM chimeric mice, 48 hours after UUO surgery, note the presence of GFP+ cells. H: Time course of fibrocyte recruitment to fibrotic kidney, spleen and full thickness skin wound. Number of cells is per sagittal section for kidney, transverse section of spleen, and transverse section of skin wound.

    Journal: The American Journal of Pathology

    Article Title: Pericytes and Perivascular Fibroblasts Are the Primary Source of Collagen-Producing Cells in Obstructive Fibrosis of the Kidney

    doi: 10.2353/ajpath.2008.080433

    Figure Lengend Snippet: Fibrocyte recruitment to kidney and spleen in response to ureteral obstruction. Male coll-GFP bone marrow was transplanted into female WT lethally irradiated mice. Chimerism was confirmed at 6 weeks by assessing leukocytes for Y chromosome. A: Image of perivascular GFP+ cells in d7 UUO kidney from coll-GFP ⇒WT bone marrow chimeric mice (magnification = original ×400). B: Confirmation of extensive fibrosis in this model by picrosirius red staining of d14 UUO kidney from chimeric mice. Note prominent perivascular fibrosis ( arrows ) as well as interstitial fibrosis. C–D: Image of d7 in UUO kidney from coll-GFP ⇒WT BM chimeric mice showing GFP+ cells co-expressing CD34 and CD45, confirming these cells to be fibrocytes (magnification = original ×400). E–F: Image of d7 UUO kidney showing BM-derived coll1a1 -GFP+ cells lacking αSMA or S100A4 expression (magnification = original ×400). G: Red pulp of spleen from coll-GFP ⇒WT BM chimeric mice, 48 hours after UUO surgery, note the presence of GFP+ cells. H: Time course of fibrocyte recruitment to fibrotic kidney, spleen and full thickness skin wound. Number of cells is per sagittal section for kidney, transverse section of spleen, and transverse section of skin wound.

    Article Snippet: Nevertheless, fibrocytes appear to constitute a novel class of myeloid dendritic cells because they lack CD11b, and express CD34.

    Techniques: Irradiation, Mouse Assay, Staining, Expressing, Derivative Assay

    Studying the effects of FGF2 on cellular calcium in satellite cells associated with host FDB fibers. Muscle cultures pre-stained with anti-CD34-FITC to identify satellite cells (A,C) were then loaded with calcium indicator X rhod-1 AM (E) . Transmitted light images are shown in (B,D) . Application of FGF2 at 2 ng/ml to the culture triggered a rise of calcium in the satellite cell in the confocal image plane ( F ; 9 satellite cells from 2 mice). Rinse with Ringer's solution without FGF2 reverses the rise in cell calcium ( G ; 9 cells from 2 mice). In cultures pre-loaded with calcium chelator BAPTA AM FGF2 caused little increase in calcium ( H ; 8 cells from 2 mice). In cultures rinsed with calcium free Ringer's solution, the effects of FGF2 are minimal ( I ; 8 cells from 2 mice). Incubation with TRPC blocker SKF 96365 attenuated the effects of FGF2 on cellular calcium ( J ; 8 satellite cells from 2 mice). FGF2 has no effects on cellular calcium of FDB fibers ( K ; 11 FDB fibers from 2 mice). ** p

    Journal: Frontiers in Physiology

    Article Title: FGF2 activates TRPC and Ca2+ signaling leading to satellite cell activation

    doi: 10.3389/fphys.2014.00038

    Figure Lengend Snippet: Studying the effects of FGF2 on cellular calcium in satellite cells associated with host FDB fibers. Muscle cultures pre-stained with anti-CD34-FITC to identify satellite cells (A,C) were then loaded with calcium indicator X rhod-1 AM (E) . Transmitted light images are shown in (B,D) . Application of FGF2 at 2 ng/ml to the culture triggered a rise of calcium in the satellite cell in the confocal image plane ( F ; 9 satellite cells from 2 mice). Rinse with Ringer's solution without FGF2 reverses the rise in cell calcium ( G ; 9 cells from 2 mice). In cultures pre-loaded with calcium chelator BAPTA AM FGF2 caused little increase in calcium ( H ; 8 cells from 2 mice). In cultures rinsed with calcium free Ringer's solution, the effects of FGF2 are minimal ( I ; 8 cells from 2 mice). Incubation with TRPC blocker SKF 96365 attenuated the effects of FGF2 on cellular calcium ( J ; 8 satellite cells from 2 mice). FGF2 has no effects on cellular calcium of FDB fibers ( K ; 11 FDB fibers from 2 mice). ** p

    Article Snippet: Primary and secondary antibodies The following primary antibodies were used: anti-laminin (L9393, rabbit IgG, Sigma, 1:200 dilution); anti-Pax-7 (mouse IgG, ascites fluid, Developmental Studies Hybridoma Bank, 1:100 dilution); anti-MyoD (M-318, rabbit IgG, Santa Cruz Biotechnology, 1:100 dilution); anti-NFATc3 (M-75, rabbit IgG, Santa Cruz Biotechnology, 1:100 dilution); anti-NFATc2 (M-300, rabbit IgG, Santa Cruz Biotechnology, 1:100 dilution); anti-CD34-FITC (RAM34, rat IgG, BD Pharmingen, 1:200 dilution); and anti-TRPC1 (T8276, rabbit IgG, Sigma, 1:100 dilution).

    Techniques: Staining, Mouse Assay, Incubation

    Identification of satellite cells on living FDB fibers. Anti-CD34-FITC antibody was added to the cell culture medium and incubated in the cell culture incubator for 3 h. After rinsing with antibody free medium, unbound antibody was washed out and only satellite cells were labeled, with negligible fluorescence appearing on host FDB fibers (A) . Culture was then fixed and immunostained with anti-Pax7 primary antibody followed by Alexa Fluor 647 conjugated secondary antibody (B) . Overlay is shown in (C) . In the transmitted light image (D) , the satellite cell is attached to the host FDB fiber.

    Journal: Frontiers in Physiology

    Article Title: FGF2 activates TRPC and Ca2+ signaling leading to satellite cell activation

    doi: 10.3389/fphys.2014.00038

    Figure Lengend Snippet: Identification of satellite cells on living FDB fibers. Anti-CD34-FITC antibody was added to the cell culture medium and incubated in the cell culture incubator for 3 h. After rinsing with antibody free medium, unbound antibody was washed out and only satellite cells were labeled, with negligible fluorescence appearing on host FDB fibers (A) . Culture was then fixed and immunostained with anti-Pax7 primary antibody followed by Alexa Fluor 647 conjugated secondary antibody (B) . Overlay is shown in (C) . In the transmitted light image (D) , the satellite cell is attached to the host FDB fiber.

    Article Snippet: Primary and secondary antibodies The following primary antibodies were used: anti-laminin (L9393, rabbit IgG, Sigma, 1:200 dilution); anti-Pax-7 (mouse IgG, ascites fluid, Developmental Studies Hybridoma Bank, 1:100 dilution); anti-MyoD (M-318, rabbit IgG, Santa Cruz Biotechnology, 1:100 dilution); anti-NFATc3 (M-75, rabbit IgG, Santa Cruz Biotechnology, 1:100 dilution); anti-NFATc2 (M-300, rabbit IgG, Santa Cruz Biotechnology, 1:100 dilution); anti-CD34-FITC (RAM34, rat IgG, BD Pharmingen, 1:200 dilution); and anti-TRPC1 (T8276, rabbit IgG, Sigma, 1:100 dilution).

    Techniques: Cell Culture, Incubation, Labeling, Fluorescence

    Repressed HIF1A expression does not significantly affect tumor growth in SCLC cell xenografts U-1906 cells, transduced with shRNA against HIF1A (sh-H1) or a non-targeting control (sh-C), were injected subcutaneously into nude mice. ( A , B ) Mean tumor specimen weight and tumor volume, 14 days after injection. A, all tumors ( n = 12) and B, specimens with confirmed tumor growth ( n = 11, see text and Panels in E). Data are presented as mean ± SEM and statistical significance was determined using Student´s t -test. Data is from one of two independent experiments ( n = 6 for each group). ( C ) Sections of formalin-fixed paraffin-embedded xenografts were characterized immunohistochemically for expression of HIF-1α, Ki67 and CD34. ( D ) The corresponding dissected tumors are shown. ( E ) Example of a specimen, devoid of tumor cells, stained with hematoxylin-eosin (H/E) and anti-Ki67 antibody. Scale bars, 100 μm.

    Journal: Oncotarget

    Article Title: Myc-induced glutaminolysis bypasses HIF-driven glycolysis in hypoxic small cell lung carcinoma cells

    doi: 10.18632/oncotarget.16904

    Figure Lengend Snippet: Repressed HIF1A expression does not significantly affect tumor growth in SCLC cell xenografts U-1906 cells, transduced with shRNA against HIF1A (sh-H1) or a non-targeting control (sh-C), were injected subcutaneously into nude mice. ( A , B ) Mean tumor specimen weight and tumor volume, 14 days after injection. A, all tumors ( n = 12) and B, specimens with confirmed tumor growth ( n = 11, see text and Panels in E). Data are presented as mean ± SEM and statistical significance was determined using Student´s t -test. Data is from one of two independent experiments ( n = 6 for each group). ( C ) Sections of formalin-fixed paraffin-embedded xenografts were characterized immunohistochemically for expression of HIF-1α, Ki67 and CD34. ( D ) The corresponding dissected tumors are shown. ( E ) Example of a specimen, devoid of tumor cells, stained with hematoxylin-eosin (H/E) and anti-Ki67 antibody. Scale bars, 100 μm.

    Article Snippet: Sections of formalin-fixed, paraffin embedded U-1906 xenograft tumors were, after antigen retrieval using PT Link (DAKO A/S, Glostrup, Denmark), stained for anti-CD34 (1:800, rat monoclonal, Santa Cruz Biotechnology, Santa Cruz, CA), anti-HIF-1α (1:50, mouse monoclonal, BD Biosciences, San José, CA) and anti-Ki67 (1:200, mouse monoclonal, DAKO A/S) using DAKO Autostainer Plus (DAKO A/S).

    Techniques: Expressing, Transduction, shRNA, Injection, Mouse Assay, Formalin-fixed Paraffin-Embedded, Staining

    In vivo principle exploration of anti-tumor using this special treatment method. a LCSM images of PANC-1 tumor slices co-stained by TUNEL immunofluorescence CD34 immunofluorescence that were harvested after sacrificing of PANC-1-bearing nude mice treated with aforementioned different treatments (G1-G5) on the 10th day post-treatment, scale bar: 50 μm, and TUNEL CD34 immunofluorescence co-staining was employed to determine cell apoptosis and vascular density, respectively, wherein FITC (green) and TRITC (red) were used to label TUNEL and CD34, respectively, and nuclei (blue) was stained by DAPI. b , c Semi-quantitative data of apoptotic cells ( b ) and apoptotic endothelial cells in all apoptotic cells ( c ) after different treatments with G1–G5 via calculating the ratios of green-labeled cells in whole blue-labeled cells and red and green co-labeled cells in all green-labeled cells according to the TUNEL immunofluorescence staining, respectively. Data are expressed as mean ± SD ( n = 3). * P

    Journal: Nature Communications

    Article Title: Extravascular gelation shrinkage-derived internal stress enables tumor starvation therapy with suppressed metastasis and recurrence

    doi: 10.1038/s41467-019-13115-3

    Figure Lengend Snippet: In vivo principle exploration of anti-tumor using this special treatment method. a LCSM images of PANC-1 tumor slices co-stained by TUNEL immunofluorescence CD34 immunofluorescence that were harvested after sacrificing of PANC-1-bearing nude mice treated with aforementioned different treatments (G1-G5) on the 10th day post-treatment, scale bar: 50 μm, and TUNEL CD34 immunofluorescence co-staining was employed to determine cell apoptosis and vascular density, respectively, wherein FITC (green) and TRITC (red) were used to label TUNEL and CD34, respectively, and nuclei (blue) was stained by DAPI. b , c Semi-quantitative data of apoptotic cells ( b ) and apoptotic endothelial cells in all apoptotic cells ( c ) after different treatments with G1–G5 via calculating the ratios of green-labeled cells in whole blue-labeled cells and red and green co-labeled cells in all green-labeled cells according to the TUNEL immunofluorescence staining, respectively. Data are expressed as mean ± SD ( n = 3). * P

    Article Snippet: In immunohistochemical staining, Anti-mouse PCNA, Abcam, cat. no. ab29; Anti-CD34 antibody, Abcam, cat. no. ab8158; Hematoxylin and Eosin Staining Kit, Beyotime Biotechnology, cat. no. C0105.

    Techniques: In Vivo, Confocal Laser Scanning Microscopy, Staining, TUNEL Assay, Immunofluorescence, Mouse Assay, Labeling

    In vivo evaluations on intratumoral vascular narrowing on PANC-1 xenografted pancreatic tumors. a CDFI images of PANC-1 xenografted pancreatic tumors implanted on nude mice that were captured at three time points, i.e., pre-, post- and 5 day post-treatment with Laser+hydrogel–GNR (2). b , c In vivo animal fluorescence images ( b ) and quantitative signal intensities ( c ) of PANC-1 xenografted tumor after corresponding treatments in control (i.e., Laser+GNR) and treated (i.e., Laser+hydrogel–GNR (2)) groups and subsequent injections of different FITC-labeled dextrans with varied sizes (i.e., 70, 20, and 4 K), wherein green arrows and dotted ellipses indicate the tumor. Data are expressed as mean ± SD ( n = 3). d , e Time-dependent ultrasonic images ( d ) and average acoustic intensities ( e ) of PANC-1 xenografted tumor before and after corresponding treatments in control and treated groups, wherein the 1st i.v. injection of Sonovue TM microbubbles were carried out, and after 30–90 min, the corresponding treatments in two groups, i.e., control (Laser+GNR) and treated (Laser+hydrogel–GNR (2)), were implemented, followed by the 2nd i.v. injection of Sonovue TM microbubbles; and the ultrasonic images were captured at three time points, i.e., pre-, post-, and 40–120 s post- each i.v. injection of Sonovue TM microbubbles, and red dotted ellipses indicate the tumor. Data are expressed as mean ± SD ( n = 3). f , g Matured ( f ) and budding ( g ) blood vessels of PANC-1 xenografted tumors after corresponding treatments in control and treated groups, and they were stained by FITC-labeled CD34 immunofluorescence ( f ) and Cy3-labeled CD31 immunofluorescence ( g ), respectively, scale bar: 200 μm. Yellow arrows indicate the intratumoral blood vessels. ( h ) In vivo animal fluorescence imaging of nude mice-bearing PANC-1 xenografted tumor at three time points, i.e., pre-, 30 s post- and 50 s post-i.t. injection of Cy3-labeled dextrans-grafted hydrogels, wherein Cy3-labeled dextran (size: 70 K) instead of GNRs was used. Statistical significances were obtained using unpaired student's t test, ** P

    Journal: Nature Communications

    Article Title: Extravascular gelation shrinkage-derived internal stress enables tumor starvation therapy with suppressed metastasis and recurrence

    doi: 10.1038/s41467-019-13115-3

    Figure Lengend Snippet: In vivo evaluations on intratumoral vascular narrowing on PANC-1 xenografted pancreatic tumors. a CDFI images of PANC-1 xenografted pancreatic tumors implanted on nude mice that were captured at three time points, i.e., pre-, post- and 5 day post-treatment with Laser+hydrogel–GNR (2). b , c In vivo animal fluorescence images ( b ) and quantitative signal intensities ( c ) of PANC-1 xenografted tumor after corresponding treatments in control (i.e., Laser+GNR) and treated (i.e., Laser+hydrogel–GNR (2)) groups and subsequent injections of different FITC-labeled dextrans with varied sizes (i.e., 70, 20, and 4 K), wherein green arrows and dotted ellipses indicate the tumor. Data are expressed as mean ± SD ( n = 3). d , e Time-dependent ultrasonic images ( d ) and average acoustic intensities ( e ) of PANC-1 xenografted tumor before and after corresponding treatments in control and treated groups, wherein the 1st i.v. injection of Sonovue TM microbubbles were carried out, and after 30–90 min, the corresponding treatments in two groups, i.e., control (Laser+GNR) and treated (Laser+hydrogel–GNR (2)), were implemented, followed by the 2nd i.v. injection of Sonovue TM microbubbles; and the ultrasonic images were captured at three time points, i.e., pre-, post-, and 40–120 s post- each i.v. injection of Sonovue TM microbubbles, and red dotted ellipses indicate the tumor. Data are expressed as mean ± SD ( n = 3). f , g Matured ( f ) and budding ( g ) blood vessels of PANC-1 xenografted tumors after corresponding treatments in control and treated groups, and they were stained by FITC-labeled CD34 immunofluorescence ( f ) and Cy3-labeled CD31 immunofluorescence ( g ), respectively, scale bar: 200 μm. Yellow arrows indicate the intratumoral blood vessels. ( h ) In vivo animal fluorescence imaging of nude mice-bearing PANC-1 xenografted tumor at three time points, i.e., pre-, 30 s post- and 50 s post-i.t. injection of Cy3-labeled dextrans-grafted hydrogels, wherein Cy3-labeled dextran (size: 70 K) instead of GNRs was used. Statistical significances were obtained using unpaired student's t test, ** P

    Article Snippet: In immunohistochemical staining, Anti-mouse PCNA, Abcam, cat. no. ab29; Anti-CD34 antibody, Abcam, cat. no. ab8158; Hematoxylin and Eosin Staining Kit, Beyotime Biotechnology, cat. no. C0105.

    Techniques: In Vivo, Mouse Assay, Fluorescence, Labeling, Injection, Staining, Immunofluorescence, Imaging

    A representative case of conjunctival melanoma in a 65-year-old female. Notes: The pigmented tumor is noted in the corneal limbs with abnormal conjunctival vessels ( A ). Histopathology of the resected tumor reveals epithelioid-type melanoma cells infiltrated beneath the conjunctival epithelium ( B ). VEGF immunoreactivity is clearly detected in the cytoplasm of tumor cells as a greenish coloration ( C ). In contrast, CD34-positive blood vessels are not detected in the tumor tissue ( D ). A bar indicates 50 µm; magnification ×40.

    Journal: Clinical Ophthalmology (Auckland, N.Z.)

    Article Title: Expression of VEGF in human conjunctival melanoma analyzed with immunohistochemistry

    doi: 10.2147/OPTH.S184193

    Figure Lengend Snippet: A representative case of conjunctival melanoma in a 65-year-old female. Notes: The pigmented tumor is noted in the corneal limbs with abnormal conjunctival vessels ( A ). Histopathology of the resected tumor reveals epithelioid-type melanoma cells infiltrated beneath the conjunctival epithelium ( B ). VEGF immunoreactivity is clearly detected in the cytoplasm of tumor cells as a greenish coloration ( C ). In contrast, CD34-positive blood vessels are not detected in the tumor tissue ( D ). A bar indicates 50 µm; magnification ×40.

    Article Snippet: Then, the sections were incubated with anti-VEGF (dilution 1:50, A-20; Santa Cruz Biotechnology Inc., Dallas, TX, USA) and anti-CD34 (dilution 1:50, M7165; Dako Japan Inc) antibodies at 4°C overnight.

    Techniques: Histopathology